Endothelial repair of posterior corneal wounds in rabbits. Influence of foetal calf serum in the culture medium

Foetal mouse brain cells were cultured as described previously [Sotelo, Gibbs, Gajdusek, Toh & Wurth (1980) Proc. Natl. Acad. Sci. U.S.A. 77, 653-657] without added insulin and without foetal calf serum after 12 days in culture. Examination by phase-contrast microscopy showed that these modifications did not appear to affect growth and development of the cells adversely. Silver impregnation of the cultures and indirect immunofluorescence following reaction with tetanus toxin showed that a high proportion of the cells resembled neurones. Analysis of concentrated culture medium by radioimmunoassay and high-pressure liquid chromatography (h.p.l.c.) revealed that the cells produced two main forms of immunoreactive insulin which differed from authentic pancreatic insulin in retention time. Immunoreactive somatostatin was also produced in culture and this was resolved into at least three forms by h.p.l.c. Immunoreactive insulin was also extracted from whole rat brain by using two published procedures. The method of Havrankova, Schmechel, Roth & Brownstein [Proc. Natl. Acad. Sci. U.S.A. (1978) 75, 5737-5741] consistently gave greater yields of insulin than did that of Eng & Yalow [Diabetes (1980) 29, 105-109] and the concentration was about three times that of plasma. The extracted insulin was further characterized by h.p.l.c. in each case and was found to behave like authentic pancreatic insulin. The production of insulin and somatostatin by foetal mouse brain cells in culture suggests that they may be a useful model system for studies of neuropeptide biosynthesis.

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Megakaryocytic Progenitors in Long Term Cultures from MDS Patients.

Blood ◽

10.1182/blood.v106.11.4924.4924 ◽

2005 ◽

Vol 106 (11) ◽

pp. 4924-4924

Author(s):

Maria Juliana Majado ◽

María I. Macizo ◽

Consuelo González-García ◽

Eduardo Salido ◽

José A. Molina ◽

...

Keyword(s):

Bone Marrow ◽

Culture Medium ◽

Horse Serum ◽

Calf Serum ◽

Methyl Cellulose ◽

Refractory Anemia ◽

Foetal Calf Serum ◽

Cell Counts ◽

Control Bone

Abstract Introduction: bone marrow long-term cultures (LTC) have been reported to produce very poor adherent stromal layer in MDS, but there are not many studies about the behaviour of the progenitor cells in supernatant culture medium. In this study we report the results of megakaryocytic progenitors (CFU-MK) in the supernatant of LTC of patients with refractory anemia (RA). Material and method: LTCs were performed to 11 RA patients and to 13 normal multiorgan donors as control. Bone marrow red cells were sedimented with 1% methyl-cellulose and seeded for LTC in 10ml flasks (Nalge Nunc International), with Iscover medium supplemented with horse serum, foetal calf serum, hydrocortisone and carbonic air, at a final concentration of 1x106 per ml. Flasks were placed in an incubator for 8 weeks. Half culture medium volume was renovated weekly, cell counts and assays of CFU-MK (Megacult C, Stem Cell Technologies), were performed. CFU-MK colonies were separated in three groups, containing 3–20 cells, 21–50 cells and more than 50 cells. Student t-test was used for statistical comparisons. Results: Are in the following table, expressed as mean + standard desviation. After the third week no colony growth was observed in normal such as in MDS. Growth of CFU-MK colonies containing more than 50 cells was higher in basal control bone marrow, no statistical differences were found in the rest of the cultures. Results CFU-MK(3–30 Cells) CFU-MK(20–50 Cells) CFU-MK (>50 Cells) CFU-MK total w: week Control basal 15.75±15.79 3.33±5.42 4.88±7.44 22.42±26.43 RA basal 7.85±8.66 0.80±1.01 0.70±0.98 9.45±9.46 t-St ns ns 0.047 ns Control w 1 3.54±4.42 0.7±1.44 0.74±1.48 4.98±6.62 RA w 1 3.29±4.52 0.29±0.93 0.25±0.67 3.82±5.65 t-St ns ns ns ns Control w 2 0.55±1.14 0.07±0.24 0.05±0.25 0.67±1.43 RA w 2 0.18±0.32 0.00±0.00 0.04±0.13 0.21±0.38 t-St ns ns ns ns Control w 3 0.18±0.53 0.00±0.00 0.00±0.00 0.18±0.53 RA w 3 0.00±0.21 0.00±0.00 0.00±0.00 0.00±0.21 t-St ns ns Discussion: No important difference was found in LTC supernatant CFU-MK in our RA patients, and this supports the idea that the stromal damage is more important, in the pathophysyology of MDS, than that of the stem cells.

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Cultivation of infective forms ofTrypanosoma congolensefrom trypanosomes in the proboscis ofGlossina morsitans

Parasitology ◽

10.1017/s0031182000041883 ◽

1981 ◽

Vol 82 (1) ◽

pp. 81-95 ◽

Cited By ~ 41

Author(s):

M. A. Gray ◽

I. Cunningham ◽

P. R. Gardiner ◽

A. M. Taylor ◽

A. G. Luckins

Keyword(s):

Culture Medium ◽

Primary Cultures ◽

Calf Serum ◽

Tsetse Flies ◽

Foetal Calf Serum ◽

Culture Vessel ◽

Minimum Essential Medium ◽

Glossina Morsitans ◽

Plastic Surface ◽

Dermal Collagen

SUMMARYTwo stocks ofTrypanosoma congolensewere established in culture at 28 °C using trypanosomes from the proboscides of infectiveGlossina morsitans. Successful primary cultures were initiated by placing an infected tsetse proboscis beside a bovine dermal collagen explant in Eagle's minimum essential medium supplemented with foetal calf serum. The trypanosomes multiplied rapidly in the medium and also gradually formed an adherent layer on the plastic surface of the culture vessel. Three primary cultures produced organisms infective for mice from 14, 20 and 35 days after initiation and thereafter continuously until days 76, 76 and 52 when they were discarded. Four attempts to initiate infective cultures using infected tsetse proboscides in medium without dermal explants were unsuccessful. When trypanosomes from primary cultures were placed in culture medium with proboscides from uninfected tsetse flies, the parasites multiplied, formed an adherent layer in the culture flasks and were seen in the proboscides within 24 h. A line of 1 stock was serially sub-passaged in this way 4 times during a period of 215 days. Infectivity titrations in mice indicated that primary and sub-passaged cultures each contained similar numbers of infective organisms. Another line of the same stock was also sub-passaged 4 times in medium alone over a period of 186 days. These sub-cultures again retained infectivity for mice, but titrations showed a decrease in infective organism production in the 4th sub-culture. Primary and sub-passaged cultures all included a variety of morphologically different developmental forms ofT. congolense, closely resembling those described in the labrum and hypopharynx ofGlossinaby previous workers. Short metacyclic-like trypanosomes and organisms with proteinaceous surface coats were present in infective cultures. Cultures were successfully re-established after cryopreservation at −196 °C and retained the ability to produce infective organisms.

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Comparison of two vitrification methods for cryopreservation of porcine embryos

Czech Journal of Animal Science ◽

10.17221/4298-cjas ◽

2011 ◽

Vol 49 (No. 5) ◽

pp. 183-189

Author(s):

J. Říha ◽

J. Vejnar

Keyword(s):

Ethylene Glycol ◽

Liquid Nitrogen ◽

Artificial Insemination ◽

Culture Medium ◽

Calf Serum ◽

Foetal Calf Serum ◽

Post Mortem ◽

Donor Insemination ◽

Viable Embryo ◽

Fresh Embryos

The aim of this study was to compare two vitrification methods of porcine perihatching blastocysts with regard to the success of transfer of these embryos to the recipients. Expanded, hatching, or hatched blastocysts were recovered post mortem from superovulated donors in 5.5 to 6.0 days after artificial insemination of donor gilts with homospermic doses. In protocol VS I, the embryos in perihatching developmental stage were equilibrated in a culture medium H-MEMD with 10% v/v of glycerol (1.37M solution of glycerol in medium) for 10 min and placed in a vitrification medium for 1.5 min max. (vitrification medium contained 50% v/v 2M sucrose in tridistilled water, 30% v/v of glycerol, and 20% v/v of foetal calf serum – FCS). Then they were dropped with micropipette and stored in liquid nitrogen vapour. For protocol VS II, we used H-MEMD culture medium supplemented with 20% v/v of FCS, 25% v/v ethylene glycol, and 25% v/v dimethyl sulphoxide (DMSO). Embryos were equilibrated for 10 min in a mixture of the vitrification medium and culture medium (1 : 1), and were kept in the vitrification medium for 1.5 minutes. Then they were dropped with micropipette and stored in liquid nitrogen vapour. Embryos were thawed by immersing the drop with the embryo in H-MEMD culture medium with 0.8M sucrose for 10 minutes. After thawing and washing in the medium with sucrose, all embryos were washed three times in a fresh medium and prepared for transfer. Recipients were synchronized either using Regumate-feeding followed by treatment with PMSG and HCG (gilts) or using piglet weaning (sows – 1st and 2nd parity). Recipients showing standing heat at the time of donor insemination were used for laparoscopic and non-surgical ET on day 5.5–6.0 of the cycle. The fraction of viable embryo vitrified under VS I or VS II protocol was 85% and 80%, compared to 95% in control fresh embryos (P > 0.05). Pregnancy of recipients was 57.3% (5/7), 67.0% (4/6) for VS I or VS II group and 42.7% (10/23) for control (P < 0.001). We can conclude on the basis of our data that both protocols for vitrification yielded similar results and can be used for cryopreservation of porcine embryos.  

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The differentiation of mouse gonads in vitro: a light and electron microscopical study

Development ◽

10.1242/dev.97.1.189 ◽

1986 ◽

Vol 97 (1) ◽

pp. 189-199

Author(s):

Sarah Mackay ◽

Robert A. Smith

Keyword(s):

Electron Microscopy ◽

Culture Medium ◽

Light And Electron Microscopy ◽

Calf Serum ◽

Microscopical Study ◽

Electron Microscopical Study ◽

The Other ◽

Foetal Calf Serum ◽

Electron Microscopical

Indifferent urogenital complexes were excised from mouse foetuses assessed by developmental criteria as day 10·5 or 11. After 4 or 6–7 days in culture, complexes were fixed and examined by light and electron microscopy. The effect of culturing sexed complexes in mixed sex groups was investigated. The effect of the presence or absence of foetal calf serum in the culture medium was considered. No evidence of inhibition of one sex by the other was found. Ovaries developed further in cultures than testes.

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The culture of mouse blastocysts in the presence of uterine flushings collected during normal pregnancy, delayed implantation and pro-oestrus

Development ◽

10.1242/dev.41.1.295 ◽

1977 ◽

Vol 41 (1) ◽

pp. 295-300

Author(s):

R. J. Aitken

Keyword(s):

Culture Medium ◽

Cell Transformation ◽

Calf Serum ◽

Normal Pregnancy ◽

Foetal Calf Serum ◽

Delayed Implantation ◽

Simple Medium ◽

Uterine Proteins ◽

Stimulation Of ◽

Blastocyst Hatching

When day-3 mouse embryos were cultured in a simple medium supplemented with uterine fluids of mice autopsied on day 4 of pregnancy, 48 h after administration of oestradiol, or during pro-oestrus, the percentage of blastocysts hatching from the zona pellucida was significantly greater than in unsupplemented medium. In the presence of uterine fluids recovered during delayed implantation this stimulation of blastocyst hatching was not observed. When the culture medium was supplemented with dialysed uterine flushings containing 20 or 30 μg protein/ml, both ‘day 4’ and ‘delay’ uterine proteins were equally effective in enhancing hatching frequency (P < 00·5). The results suggested that ‘delay’ uterine fluids may contain a dialysable inhibitor of blastocyst activity. The putative inhibitor was not effective in the presence of serum, since uterine fluids recovered both on day 4 of pregnancy and during delayed implantation significantly increased the size attained by lastocyst outgrowths in the presence of foetal calf serum (P < 0·001). The percentage of blastocysts exhibiting giant cell transformation and outgrowth was also increased (P < 0·02) by these uterine fluids when the concentration of FCS in the medium was minimal (0·25 %).

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TESTOSTERONE METABOLISM IN CULTURED HYPERPLASIA OF THE HUMAN PROSTATE

Acta Endocrinologica ◽

10.1530/acta.0.0770784 ◽

1974 ◽

Vol 77 (4) ◽

pp. 784-793 ◽

Cited By ~ 10

Author(s):

M. J. McMahon ◽

A. V. J. Butler ◽

G. H. Thomas

Keyword(s):

Benign Prostatic Hyperplasia ◽

Organ Culture ◽

Culture Medium ◽

Calf Serum ◽

Culture Conditions ◽

Prostatic Hyperplasia ◽

Human Prostate ◽

Foetal Calf Serum ◽

Testosterone Metabolism ◽

Culture Time

ABSTRACT Slices of human benign prostatic hyperplasia were maintained in organ culture in the presence of [3H]- or [14C]testosterone. Explants concentrated radioactivity from the culture medium, although this effect was depressed by the inclusion of foetal calf serum in the medium. Testosterone was metabolised to products with chromatographic mobilities corresponding to androstanediols, androsterone, dehydroepiandrosterone, and androstanedione. The principal metabolite was identified as 5α-dihydrotestosterone, and small amounts of testosterone and androstenedione were also found. As culture time increased from 1 to 6 days there was a diminution in the proportion of 5α-, and an increase in 17-ketometabolites, indicating a swing to a less physiological pattern of testosterone metabolism by the aging culture. Testosterone metabolism was thought to provide a sensitive index by which to evaluate changes in culture conditions.

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The Effects of Hyaluronic Acid, Calcium Hydroxide, and Dentin Adhesive on Rat Odontoblasts and Fibroblasts

Archives of Industrial Hygiene and Toxicology ◽

10.2478/10004-1254-62-2011-2076 ◽

2011 ◽

Vol 62 (2) ◽

pp. 155-161 ◽

Cited By ~ 8

Author(s):

Ana Bogović ◽

Jana Nižetić ◽

Nada Galić ◽

Davor Želježić ◽

Vedran Micek ◽

...

Keyword(s):

Hyaluronic Acid ◽

Calcium Hydroxide ◽

Culture Medium ◽

Calf Serum ◽

Foetal Calf Serum ◽

Sprague Dawley ◽

Pulp Tissue ◽

Pulp Capping ◽

Sprague Dawley Rats ◽

Dentin Adhesive

The Effects of Hyaluronic Acid, Calcium Hydroxide, and Dentin Adhesive on Rat Odontoblasts and FibroblastsThe aim of this study was to investigate the effects and efficiency of pulp capping preparations based on hyaluronic acid, calcium hydroxide, and dentin adhesive on the pulp tissue of Sprague-Dawley rats. The rats were killed and extracted teeth sectioned transversely through the pulp. The slices were placed in a RPMI 1640 cell culture medium supplemented with 10 % foetal calf serum. During 14 days of cultivation cultures were treated with preparations that contained hyaluronic acid (Gengigel Prof®), and calcium hydroxide (ApexCal®), or with dentin adhesive (Excite®). Cellularity and viability of fibroblasts and odontoblasts was analysed using a haemocytometer. Hyaluronic acid proved most efficient and the least toxic for direct pulp capping. Even though calcium hydroxide and dentin adhesive demonstrated a higher degree of cytotoxicity, their effects were still acceptable in terms of biocompatibility.

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