scholarly journals In vitro mouse spermatogenesis with an organ culture method in chemically defined medium

PLoS ONE ◽  
2018 ◽  
Vol 13 (2) ◽  
pp. e0192884 ◽  
Author(s):  
Hiroyuki Sanjo ◽  
Mitsuru Komeya ◽  
Takuya Sato ◽  
Takeru Abe ◽  
Kumiko Katagiri ◽  
...  
Keyword(s):  
Organ Culture ◽  
Culture Method ◽  
Defined Medium ◽  
Chemically Defined Medium

Accelerating effect of hydrocortisone on the keratinization of chick embryonic skin growing in a chemically defined medium

Development ◽  
1971 ◽  
Vol 25 (3) ◽  
pp. 365-376
Author(s):  
Masanobu Sugimoto ◽  
Hiroyoshi Endo
Keyword(s):  
Culture Method ◽  
Defined Medium ◽  
Chemically Defined Medium ◽  
In Ovo ◽  
Millipore Filter ◽  
Embryonic Skin ◽  
Minimal Concentration ◽  
Chick Embryonic Skin

In an attempt to examine histologically, chemically and biochemically the effect of hydrocortisone in a minimal concentration on keratinization of 13-day chick embryonic shank skin, a simple replicate culture method (‘Millipore’ filter-roller-tube method) was devised to cultivate rather large pieces of the skin in a chemically defined medium, BGJb supplemented with ascorbate. Hydrocortisone added in a minimal concentration of 0·001 μg/ml produced a heavily cornified eosinophilic layer over the epidermis after 4 days' cultivation, whereas in the absence of the steroid no sign of cornification could be found during culture. Determination of total protein and analysis of amino acid composition of whole protein of the epidermis indicated that hydrocortisone accelerated epidermal cornification as compared with in ovo development. Pregnenolone and progesterone showed no effect on the in vitro keratinization of the epidermis and deoxycorticosterone gave a slight effect: thus the cornification-accelerating effect of hydrocortisone seems to be attributable to its glucocorticoid activity.

Organ Culture of the Mammalian and Avian Embryo Otocyst

1977 ◽  
Vol 86 (3) ◽  
pp. 371-380 ◽  
Author(s):  
I. Friedmann ◽  
M. Giselle Hodges ◽  
P. N. Riddle
Keyword(s):  
Organ Culture ◽  
Inner Ear ◽  
Middle Ear ◽  
Defined Medium ◽  
Avian Embryo ◽  
Vestibular Apparatus ◽  
Chemically Defined Medium ◽  
Organ Cultures ◽  
Cochlear Duct

The use of a chemically defined medium supplemented with serum has proved most suitable for the growth of the isolated embryonic otocyst in vitro. Complete differentiation of the cochlear duct and of the vestibular apparatus ensued and is described. The system is reliable for organ cultures of both the avian and mammalian otocyst and for the study of various lesions affecting the inner ear and middle ear.

A Chemically Defined Medium Supports in Vitro Proliferation and Maintains the Osteochondral Potential of Rat Marrow-Derived Mesenchymal Stem Cells

1995 ◽  
Vol 219 (1) ◽  
pp. 211-222 ◽  
Author(s):  
Donald P. Lennon ◽  
Stephen E. Haynesworth ◽  
Randell G. Young ◽  
James E. Dennis ◽  
Arnold I. Caplan
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Defined Medium ◽  
Chemically Defined Medium ◽  
In Vitro Proliferation

197 EFFECTS OF HYALURONAN ON OXYGEN CONSUMPTION AND ATP CONTENT IN PIG BLASTOCYSTS PRODUCED IN VITRO

2007 ◽  
Vol 19 (1) ◽  
pp. 215
Author(s):  
K. Yoshioka ◽  
M. Yokoo ◽  
T. Ozawa ◽  
C. Suzuki ◽  
H. Abe ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Metabolic Activity ◽  
Formation Rate ◽  
Defined Medium ◽  
Cell Number ◽  
Total Cell ◽  
Chemically Defined Medium ◽  
Atp Content ◽  
Cell Numbers

Hyaluronan (HA), a glycosaminoglycan present in follicular and oviductal fluids, has been related to sperm capacitation, fertilization, and embryo development. We have found that exogenous HA improves cell proliferation of porcine embryos cultured in a chemically defined medium (Yoshioka et al. 2004 Reprod. Fertil. Dev. 16, 264–265). Moreover, mitochondrial maturation was clearly more advanced in blastocysts cultured with HA compared to those cultured without HA, as seen by transmission electron microscopy. In the present study, the effects of HA on oxygen consumption and ATP content of blastocysts, produced in a defined system which reflects metabolic activity, were investigated. Porcine immature oocytes were matured for 44 h in porcine oocyte medium (POM) and subsequently fertilized with frozen–thawed ejaculated semen in porcine gamete medium supplemented with theophylline, adenosine, and cysteine (PGMtac4). Both POM and PGMtac4 were chemically defined media modified from porcine zygote medium (PZM)-5. After IVF, presumptive zygotes were cultured in PZM-5 containing HA (from the microorganism, Nacalai tesque, Kyoto, Japan) at concentrations of 0 [control], 10 [HA10], or 100 [HA100] �g mL-1 until 5 days after IVF. Blastocyst formation rate and total cell numbers/blastocyst at Day 5 were assessed. In addition, oxygen consumption and ATP content of single Day 5 blastocysts were measured. Blastocyst oxygen consumption was quantified using scanning electrochemical microscopy (HV-403; Research Institute for the Functional Peptides, Yamagata, Japan), and embryonic ATP content was determined using a commercial assay based on the luciferin-luciferase reaction (ATPlite; PerkinElmer, Groningen, The Netherlands). Data were statistically analyzed by ANOVA and Fisher's PLSD test. While the percentage of embryos that developed to the blastocyst stage [30.5% (63/206) to 31.7% (65/206)] did not differ among treatments, blastocyst cell number in the HA100 group [57.9 cells (n = 64)] was greater (P < 0.05) compared to those in the control [48.6 cells (n = 63)] or HA10 [50.0 cells (n = 65)] groups. Blastocyst oxygen consumption rate in the HA100 group [0.629 � 10-14 mol s-1 (n = 15)] was significantly higher than in the control [0.500 � 1-14 mol s-1 (n = 16)] or HA10 [0.464 � 10-14 mol s-1 (n = 14)] groups. ATP content/blastocyst did not differ among treatments [control: 0.645 pmol (n = 38), HA10: 0.727 pmol (n = 42), and HA100: 0.704 pmol (n = 43)]. It is concluded that HA affects the metabolic activity of pig blastocysts developed in a chemically defined medium, enhancing oxygen consumption and their total cell numbers, thus improving the quality of IVP blastocysts.

224 SILDENAFIL ACCELERATES SPERM PENETRATION OF PORCINE OOCYTES IN A CHEMICALLY DEFINED MEDIUM

2013 ◽  
Vol 25 (1) ◽  
pp. 260
Author(s):  
H. Funahashi ◽  
Q.-S. Wu
Keyword(s):  
Adenosine Monophosphate ◽  
Defined Medium ◽  
Fluorescence Assay ◽  
Chemically Defined Medium ◽  
Sperm Capacitation ◽  
Boar Sperm ◽  
Sperm Penetration ◽  
Type 3 ◽  
Chlortetracycline Fluorescence Assay

Sperm capacitation, a cyclic-adenosine monophosphate-dependent phenomenon, is an important initiation step for penetration into oocytes. In porcine IVF, the use of caffeine, as a nonspecific phosphodiesterase (PDE) inhibitor, is common to accelerate sperm capacitation and penetration. The objective of this study was to examine the effects of PDE inhibitors (cilostamide, rolipram, and sildenafil as PDE type 3-, type 4-, and type 5-specific inhibitors, respectively) on the capacitation of boar sperm and the penetration into porcine oocytes in the absence of caffeine and other capacitation inducers in a chemically defined medium. After washing sperm samples collected from an ejaculated sperm-rich fraction of different individual Berkshires, the sperm were resuspended in capacitation inducer-free (theophylline- and adenosine-free) PGM-tac4 (mPGM-tac) at 5 × 105 cells mL–1. The suspension was cultured in mPGM-tac nonsupplemented or supplemented with 2.5 mM cilostamide, rolipram, or sildenafil for 90 min at 39°C in an atmosphere of 5% CO2 in air, and then the capacitation status was assessed by chlortetracycline fluorescence assay. Other sperm suspensions were used to co-culture with denuded in vitro-matured oocytes in the same medium for 8 h in an atmosphere of 5% CO2 in air and fixed, and sperm penetration was then examined. Statistical analyses of results from 4 replicated trials were performed by ANOVA with a Bonferroni-Dunn post hoc test (significance, P < 0.05). In our result from the chlortetracycline fluorescence assay, although the incidence of intact sperm was significantly reduced in the presence of rolipram (54.3%) and sildenafil (52.7%) as compared with controls (66.7%), there were no differences in capacitated sperm among experimental groups (24.3 to 34.3%). The incidence of acrosome-reacted sperm was higher in the presence of cilostamide (17.3%) than in the others (9.0 to 13.0%). High sperm penetration was observed only in the presence of sildenafil (76.6%) as compared with the control (0%) or the presence of rolipram (4.4%) or cilostamide (1.8%). These results demonstrate that inhibition of PDE type 5, but not PDE type 3 and type 4, significantly accelerates the penetration of boar sperm into the oocytes in a capacitation inducer-free chemically defined medium, whereas inhibition of PDE type 3 may induce an acrosome reaction.

Schistosoma mansoni: evidence for a role of serum factors in protecting artificially transformed schistosomula against antibody-mediated killing in vitro

Parasitology ◽  
1978 ◽  
Vol 77 (2) ◽  
pp. 225-233 ◽  
Author(s):  
C. A. P. Tavares ◽  
Rita C. Soares ◽  
P. M. Z. Coelho ◽  
G. Gazzinelli
Keyword(s):  
Schistosoma Mansoni ◽  
Protective Factor ◽  
Gel Filtration ◽  
Defined Medium ◽  
Rabbit Serum ◽  
Chemically Defined Medium ◽  
Lethal Effects ◽  
Serum Factors

SummaryArtificially transformed schistosomula of Schistosoma mansoni develop a consistent but small protection against the lethal effects of antibody plus complement when cultured for 24 h in a chemically defined medium. In contrast, they become rapidly resistant to antibody plus complement, when cultured in the presence of a complex medium consisting of equal parts of heat-inactivated rabbit serum and Earle's/lactalbumin or in defined medium supplemented with small amounts of heat-inactivated rabbit serum. Sephadex G-200 gel filtration revealed that the protective factor in rabbit serum is a macromolecule with a molecular weight between 7 and 19S. Parasites cultured at 10 °C or in the presence of 200 μg of puromycin show less serum-induced protection against the lethal effects of antibody plus complement than do controls.

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