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PubMed1992

Evaluating an in vitro culture system of bovine uterine and oviduct epithelial cells for subsequent embryo co-culture.

Three experiments were conducted to evaluate the effects of culture medium and incubation temperature on bovine uterine and oviduct epithelial cell growth, so that the most efficient combination could then be used to develop a co-culture system for bovine embryos. In the first experiment, uterine and oviduct epithelial cells at either the second or third subpassage were incubated for 8 days at 37 degrees C with 5% CO2 in Tissue Culture Medium-199, CMRL-1066, Minimal Essential Medium, Ménézo's B2 or Ham's F-12 medium. In addition to plotting growth curves of cell populations, the cell cycle was monitored for 8 days by flow cytometry. Uterine and oviduct epithelial cells incubated in CMRL-1066 exhibited the highest growth rates during the 8-day culture period. However, there were no differences in cell cycle analysis among treatment groups during the incubation period. In the second experiment, CMRL-1066 medium was used to evaluate growth and proliferation of uterine and oviduct epithelial cells incubated at 37 degrees C or 39 degrees C; temperature had no significant effect on growth rates or proliferation rates for either uterine or oviduct cells during the 8-day incubation. In the third experiment, the more promising culture media for epithelial cell culture studies were chosen for in vitro maturation and subsequent in vitro fertilization (IVF) of bovine oocytes. Early cleavage-stage embryos produced by IVF procedures were subsequently cultured in vitro for 7 days in medium alone or with oviduct epithelial cells. In this study, the culture medium did not influence fertilization or cleavage rates. However, more embryos co-cultured with oviduct epithelial cells were considered viable after 7 days of incubation compared with embryos incubated in medium alone. These results indicate that various incubation conditions can influence the growth of bovine uterine and oviduct epithelial cells in vitro. However, in spite of changes in cell growth patterns, there does not appear to be a change in their embryotropic capabilities in vitro.

Animals↗
PubMed1994

Enhanced rates of cleavage and development for sheep zygotes cultured to the blastocyst stage in vitro in the absence of serum and somatic cells: amino acids, vitamins, and culturing embryos in groups stimulate development.

The aim of this study was to develop a serum-free culture system that could support high levels of cleavage and blastocyst formation from sheep zygotes developed in vitro. To this end, we investigated the effects on sheep zygote development of amino acids, ammonium, vitamins, and culture of embryos in groups in Synthetic Oviduct Fluid (SOF) medium supplemented with BSA (32 mg/ml). The inclusion of amino acids in the culture medium had no effect on the percentage of embryos arrested at the 8-16-cell stage when embryos were cultured singly in the same drop of medium for 6 days (43% in SOF; 41% in SOF+amino acids). However, in medium containing all Eagle's amino acids, replacing the culture medium every 48 h to alleviate ammonium toxicity significantly decreased the number of arrested embryos (6%; p < 0.05) and significantly increased blastocyst cell number (52 cells in SOF; 105 cells in SOF+amino acids; p < 0.01) and the number of embryos developing to the blastocyst stage (29% in SOF; 67% in SOF+amino acids; p < 0.05). When the medium was renewed every 48 h, nonessential amino acids and glutamine also significantly decreased the number of arrested embryos (p < 0.05). Culturing embryos singly or in groups in SOF medium with all Eagle's amino acids that was renewed every 48 h resulted in significant increases in blastocyst hatching and mean cell number (47%, 31%, and 79%; 105, 136, and 173 cells for embryos cultured singly, in groups of 2, and in groups of 4, respectively). After culture in groups of 4, blastocyst cell numbers were equivalent to in vivo-developed controls (160 cells) and significantly greater than those developed in serum (103 cells; p < 0.01). Analysis of blastocyst metabolism, expressed on a per-cell basis, revealed that amino acids did not affect either glucose uptake or lactate production, whereas the addition of amino acids and vitamins resulted in a significant increase in both parameters (p < 0.01). A similar response was observed in serum-derived blastocysts. Ammonium production by sheep blastocysts after culture in the presence of amino acids was significantly greater than that produced by mouse blastocysts, indirect evidence that ruminant embryos utilize amino acids to a greater extent than do rodent embryos.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acids↗
PubMed1999

Developmental competence and metabolism of bovine embryos cultured in semi-defined and defined culture media.

Development of in vitro-produced bovine embryos was studied in 3 two-step culture media: synthetic oviduct fluid (SOF), Gardner's G1/G2, and control (hamster embryo culture medium with 11 amino acids [HECM-6] followed by tissue culture medium 199 + 10% bovine calf serum). Modifications were made to reduce or eliminate protein. Glycolysis and Krebs cycle activity of morulae and blastocysts developed from selected immature oocytes were measured. There were no differences in development to the morula and blastocyst stages between SOF, G1/G2, or control (41%, 36%, and 46%, respectively), although more blastocysts developed in control medium than in G1/G2 (46%, 30%, respectively). Reducing or removing BSA during the initial culture period did not significantly reduce development to blastocyst (31%, 33%, respectively), although development was reduced in SOF with BSA removed from the final culture period (19%). There were no differences in development to the blastocyst stage between SOF, SOF with BSA removed during the initial culture period, and control (44%, 32%, 49%, respectively), but development was reduced in chemically defined protein-free medium throughout the culture period (21%). Krebs cycle activity did not differ between treatments; however, glycolysis was highest in the control embryos and lowest in embryos cultured in protein-free medium. Embryos that developed in the presence of serum appeared dark and granular and had elevated glycolytic rates compared to embryos developed in completely defined medium. This study shows that both metabolism and blastocyst development of embryos are altered by different culture media, implying a functional linkage between these two indicators of successful embryogenesis.

Animals↗
PubMed2005

Considering the culture of disability in cultural competence education.

Cultural competence extends beyond understanding those values, beliefs, and needs that are associated with patients' age or gender or with their racial, ethnic, or religious backgrounds. People hold many simultaneous cultural associations, and each have implications for the care process. The "culture of disability" is a pan-ethnic culture for which a set of physician competencies are required to ensure appropriate, culturally sensitive care to persons with congenital or acquired disabilities. Such competencies include communicating with patients who have deficits in verbal communication and avoidance of infantilizing speech; understanding the values and needs of persons with disabilities; the ability to encourage self-advocacy skills of patients and families; acknowledging the core values of disability culture including the emphasis on interdependence rather than independence; and feeling comfortable with patients with complex disabilities. Medical schools have developed programs to increase students' exposure to persons with disabilities and it is suggested that such programs are most effective when they are the result of collaboration with community-based facilities or organizations that serve persons with disabilities in the natural environment. Combining lecture-based instruction and structured experiences with the opportunity for students to interact with patients in their natural environments may facilitate development of competencies with respect to patients with disabilities. The culture of disability should be included as one of the many cultures addressed in cultural competence initiatives in medical school and residency curricula.

Attitude to Health↗
PubMed2005

Reorganization of hair follicles in human skin organ culture induced by cultured human follicle-derived cells.

Studies of human hair follicle (HF) induction by follicle-derived cells have been limited due to a lack of suitable test systems. In this study, we established a skin organ culture system which supports HF formation by follicle-derived cells. Long-term skin organ cultures were set up from human retroauricular skin specimens and maintained in culture for up to 8 weeks. In vitro expanded human HF-derived cells from the dermal papilla (DP) and the outer root sheath (ORS) were injected together into the skin specimens and evaluated for their ability to induce reorganization of HFs. Macroscopic analysis of the cultured skin specimens demonstrated the growth of velus-like hair after 4 weeks in culture. Histologic evaluation of the cultured skin specimens after 8 weeks of culture revealed multiple miniaturized HFs with sebaceous glands. In addition, cell clusters of various differentiation stages could be demonstrated in serial sections of the cultured skin specimens. Labeling of HF-derived cells with the fluorescence dye CFDA-1 prior to injection suggested a de novo reorganization of HFs out of the injected cells. In conclusion, the study demonstrated HF formation by HF-derived cells in an in vitro skin organ culture model.

Adult↗
PubMed1975

Organ culture of foetal rat pancreas. Effects of parabiotic culture with foetal adrenal glands.

The differentiation and growth of the foetal rat pancreas (20 days postcoitum) was studied in parabiotic organ culture with foetal adrenal tissue. In such co-cultures, characteristic pancreatic morphology was preserved and further acinar cell differentiation was fostered. Acinar cells contiunued to represent about 65% of the total explant volume following short-term incubation. The selective islet cell proliferation, previously observed in control pancreatic explants cultured alone, did not occur when adrenals were co-cultured. In addition, the amylase content of the incubation media and of the explanted pancreatic tissue remained high with adrenal co-culture, while the insulin content of the media and of the explanted tissue was markedly suppressed when compared to control pancreatic explants cultured alone. The effects of the adrenal in maintaining the differentiated acinar component of the pancreas and suppressing media insulin concentration diminished over extended incubation. The addition of adrenals to culture of foetal pancreatic explants after 6 days of control culture (at a time when differentiated acinar cells were not identifiable in the explant) did not result in redifferentiation of the acinar component, but did markedly depress media insulin content. Removal of adrenals after 4 days of co-culture resulted in an immediate rise in media insulin concentration and a rapid decline in pancreatic acinar mass. An adrenal-exocrine pancreatic axis is proposed and it is suggested that foetal adrenal secretions may play an important role in the development of the exocrine pancreas in vivo as well as in vitro.

Adrenal Cortex Hormones↗
PubMedDate not supplied

Organotypic brain-slice cultures from adult rats: approaches for a prolonged culture time.

Animal experiments are widely used in neurobiological and neuropharmacological research. Today, juvenile brain organotypic slice cultures have partially replaced in vivo experiments, but there is no adequate in vitro counterpart for the adult brain. The present study was aimed at the long-term culture of physiologically intact hippocampal slices from adult rats, by improving the conditions for preparation and culture, and the development of a new culture medium. A cerebrospinal fluid (CSF)-like medium was used, which was modified with a variety of supplements, including energy precursors, free-radical scavengers, and compounds known to inhibit neurotoxicity. The population spike amplitude (PSA) was used as a measure of viability, and amplitudes larger than 1mV indicated viable cultures. The addition of MK-801 during slice preparation improved PSA values during the first two days in vitro (DIV). Ascorbic acid and insulin prolonged the culture time up to DIV 4. FK-506 and vitamin E, alone or in combination, supported slice culture up to DIV 5. An increase in ATP, unless combined with vitamin E, and/or insulin, increased culture time up to DIV 6. Vitamins B(1), B(2), B(12) and D(2) had no effect. The modified CSF-like medium developed in this study permits the culture of adult hippocampal tissue for at least 6 days.

Animals↗
PubMed1987

Anthropology and psychiatry. The role of culture in cross-cultural research on illness.

To illustrate the contribution anthropology can make to cross-cultural and international research in psychiatry, four questions have been put to the cross-cultural research literature and discussed from an anthropological point of view: 'To what extent do psychiatric disorders differ in different societies?' 'Does the tacit model of pathogenicity/pathoplasticity exaggerate the biological aspects of cross-cultural findings and blur their cultural dimensions?' 'What is the place of translation in cross-cultural studies?' and 'Does the standard format for conducting cross-cultural studies in psychiatry create a category fallacy?' Anthropology contributes to each of these concerns an insistence that the problem of cross-cultural validity be given the same attention as the question of reliability, that the concept of culture be operationalised as a research variable, and that cultural analysis be applied to psychiatry's own taxonomies and methods rather than just to indigenous illness beliefs of native populations.

Anthropology, Cultural↗
PubMed1983

Growth hormone secretion by cultured rat anterior pituitary cells. Effects of culture conditions and dexamethasone.

Optimal conditions were sought for the study of GH secretion by cultured normal pituitary cells. Dispersed rat pituitary cells were cultured for 1 week in four different media supplemented with 10% fetal calf serum. Minimal essential medium resulted in high GH content and secretion during a 4-h incubation period, whereas GH secretion was lower (P less than 0.05) for cells cultured in medium 199, Ham's F-10, and RPMI-1640. GH secretion/24 h declined gradually with time. After 2 weeks in culture hormone secretion amounted to 30% of secretion on day 1, but after 3 weeks GH secretion was still measurable. GH recovery during the 3-weeks culture period was more than 600% of the amount initially plated. GH secretion was positively correlated with the bicarbonate concentration between 0.85 and 2.2 g/liter NaHCO3. When pituitary cells were cultured in concentrations varying from 0.5 X 10(5) to 10 X 10(5) cells per dish, GH secretion and content per cell were constant, suggesting that no direct autofeedback occurred in cultures with high cell densities and thus high medium GH. Dexamethasone stimulated GH secretion and content in a dose-dependent way (0.1 nM-10 microM). The stimulatory effect of 100 nM dexamethasone occurred within 24-48 h. After 7 days of treatment with 100 nM dexamethasone, GH secretion had increased to 190% and GH content to 230% of control. In contrast to the effects on GH, dexamethasone suppressed PRL secretion in a dose-dependent way, but this effect was seen only after 7 days of treatment and not after 4 days of treatment. Cycloheximide and actinomycin D prevented the stimulatory effect of dexamethasone on GH secretion. However, 24 h after cessation of cycloheximide treatment GH secretion was stimulated by dexamethasone. Four days of treatment with 100 nM dexamethasone did not affect the GH response to somatostatin, prostaglandin E1, and theophylline, nor the PRL response to dopamine, TRH, and theophylline. Thus, culture conditions may affect GH production, and dexamethasone can be used to culture somatotrophs for longer periods with steady GH production and normal responsiveness.

Alprostadil↗
PubMed2001

Culture and eating disorders: a historical and cross-cultural review.

Cultural beliefs and attitudes have been identified as significant contributing factors in the development of eating disorders. Rates of these disorders appear to vary among different racial/ethnic and national groups, and they also change across time as cultures evolve. Eating disorders are, in fact, more prevalent within various cultural groups than previously recognized, both within American ethnic minorities and those in other countries. This review examines evidence for the role of culture as an etiological factor for the development of eating disorders. Historical and cross-cultural experiences suggest that cultural change itself may be associated with increased vulnerability to eating disorders, especially when values about physical aesthetics are involved. Such change may occur across time within a given society, or on an individual level, as when an immigrant moves into a new culture. Further research into the cultural factors that promote the development of eating disorders is much needed. Understanding how cultural forces contribute to the development of disorders is needed so that preventive interventions can be created.

Body Image↗
PubMed1989

Effects of various culture conditions on matrix formative functions of rat incisor odontoblasts in a pulp-dentin slice culture system.

An attempt to develop a new pulp-dentin slice culture system was carried out using thin slices of rat incisors in order to obtain a better culture method which will be useful for studies of the physiological function of odontoblasts including the mechanism of dentiogenesis. The thin slices of incisors were prepared from mandibles of 4-5-day-old rats, using original equipment that was developed in the present study. They were incubated under stationary and rocking culture conditions with two oxygen tensions (50 and 95%). The hyperbaric condition (30% O2, 2 atm) was also tested. Histological observations, [3H]-proline uptake examination and [3H]-proline autoradiography were carried out in order to estimate the matrix formative ability of odontoblasts. The results showed that the pulp-dentin slice culture system successfully preserved the viability and the matrix formative function of odontoblasts. The rocking culture was more favorable than the stationary culture to maintain the cell viability. The higher oxygen tension improved the matrix formative function of odontoblasts. Satisfactory results were obtained from the rocking culture (95% O2) and the hyperbaric stationary culture (30% O2, 2 atm). These results suggest that a large amount of oxygen may be required for maintaining the functions of odontoblasts in this new pulp-dentin slice culture system.

Animals↗
PubMed1989

Toxic effects of 7 beta-hydroxycholesterol on rat liver primary cultures, epithelial lines and co-cultures.

The toxic effects of 7 beta-hydroxycholesterol (7 beta-OHC) on cultures and co-cultures of rat hepatocytes, rat liver epithelial cell lines, and rat liver fibroblast lines were investigated. Hepatocytes in primary culture or co-cultured with proliferative epithelial cells, were not affected by the presence of 7 beta-OHC at a concentration of 400 microM over a period of 72 hours. In contrast, proliferative cultures of liver epithelial cell lines and liver fibroblast lines were killed by 50 microM 7 beta-OHC within the first 24 hours. Established liver epithelial cells (hyperploid) were more sensitive to 7 beta-OHC than the same line at early passages (diploid). When hepatocytes and liver epithelial cells were co-cultured and treated with 100 microM 7 beta-OHC, only epithelial cells were lysed. A concentration of 50 microM 7 beta-OHC was toxic to co-cultures of liver epithelial cell and fibroblasts together. In a serum-free medium, the cytotoxic concentration of 7 beta-OHC was lower than that in the serum-supplemented medium. Thus, liver epithelial cells cultured alone or co-cultured with hepatocytes were killed at 12.5 microM and 50 microM 7 beta-OHC, respectively. Finally, cholesterol concentrations four-fold that of 7 beta-OHC antagonized the lethal effects of 7 beta-OHC in the serum-free medium.

Albumins↗
PubMed2000

Differentiation of mammalian skeletal muscle cells cultured on microcarrier beads in a rotating cell culture system.

The growth and repair of adult skeletal muscle are due in part to activation of muscle precursor cells, commonly known as satellite cells or myoblasts. These cells are responsive to a variety of environmental cues, including mechanical stimuli. The overall goal of the research is to examine the role of mechanical signalling mechanisms in muscle growth and plasticity through utilisation of cell culture systems where other potential signalling pathways (i.e. chemical and electrical stimuli) are controlled. To explore the effects of decreased mechanical loading on muscle differentiation, mammalian myoblasts are cultured in a bioreactor (rotating cell culture system), a model that has been utilised to simulate microgravity. C2C12 murine myoblasts are cultured on microcarrier beads in a bioreactor and followed throughout differentiation as they form a network of multinucleated myotubes. In comparison with three-dimensional control cultures that consist of myoblasts cultured on microcarrier beads in teflon bags, myoblasts cultured in the bioreactor exhibit an attenuation in differentiation. This is demonstrated by reduced immunohistochemical staining for myogenin and alpha-actinin. Western analysis shows a decrease, in bioreactor cultures compared with control cultures, in levels of the contractile proteins myosin (47% decrease, p < 0.01) and tropomyosin (63% decrease, p < 0.01). Hydrodynamic measurements indicate that the decrease in differentiation may be due, at least in part, to fluid stresses acting on the myotubes. In addition, constraints on aggregate size imposed by the action of fluid forces in the bioreactor affect differentiation. These results may have implications for muscle growth and repair during spaceflight.

Animals↗
PubMed1986

Effect of culture age on drug metabolizing enzymes and their induction in primary cultures of rat hepatocytes.

In order to further establish optimal and reproducible conditions for the use of primary hepatocyte cultures in studies of drug metabolism, the effect of culture age on the basal and induced activities of ethoxycoumarin O-deethylase (ECDE), UDP-glucuronyltransferase (GT) [methylumbelliferone (MU) and p-nitrophenol (pNP) as substrates] and sulfotransferase (MU) were measured. In contrast to the monooxygenase activity conjugating activities were maintained for 2-3 weeks in culture, although especially sulfate conjugation showed a transient decline during the first days, and GT activity increased later on during culture. Low induction of ECDE with both phenobarbital (PB) and 3-methylcholanthrene (MC) was seen during the first day in culture, and maximum induction was obtained when inducer was added on the second or third day. The MC inducible GT (pNP) exhibited a similar behaviour indicating that the coordinated induction of the MC inducible activities is preserved in culture. The results show that primary cultures of hepatocytes can be used to study conjugating enzymes and their regulation. However, each functional parameter that is to be investigated in hepatocyte cultures should first be studied as a function of culture age to establish the optimum time.

7-Alkoxycoumarin O-Dealkylase↗
PubMed1985

Selective culture of neurons from rat cerebral cortex: morphological characterization, glutamate uptake and related enzymes during maturation in various culture media.

Dissociated cerebral cortex of fetal rat was grown in a serum-free, chemically defined medium (CDM) (containing insulin, progesterone, estradiol, transferrin, putrescine, selenium and 15 mM KCl) and compared with cultures grown in a medium containing 20% fetal calf serum (SCM). Neurons survived well using either medium, but in the serum-free medium the cellular population was exclusively neuronal (at 96%), while glial cells began to proliferate after one week in the SCM. The various cellular morphologies are described in the present report and the presence of immunological markers characteristic of neurons was investigated. Autoradiographic experiments have been performed after incubation with various putative neurotransmitters and we have shown the presence of a strikingly high proportion of glutamatergic neurons in these cultures. Glutamate high affinity uptake was also greatly increased in neuronal cultures maintained in a CDM compared to a SCM, especially in young cultures. The development of different enzymes involved in the metabolism of glutamate was also studied; the specific activity of glutaminase increased in culture and was found to be higher in a CDM than in a SCM, while the inverse was true for glutamine synthetase. The relative proportion of both enzymes in neurons compared to glial cells was opposite, as neuronal cultures had higher levels of glutaminase and glial cultures were enriched in glutamine synthetase activity. It seems that the proportion of glutamate-neurons increases when cultured in a CDM compared to a SCM and we suggest that this culture procedure may provide a purely neuronal population enriched in mature glutamatergic neurons. It may thus be useful for future in vitro studies on glutamate and GABA metabolism in neurons.

Animals↗
PubMed1992

'Culture' in culture-bound syndromes: the case of anorexia nervosa.

Anorexia nervosa is presently considered a Western culture-bound syndrome. A cultural focus on dieting and ideals of thinness for women are assumed to be implicated in the disorder. While research indicates that the majority of non-anorectic women in the United States are preoccupied with body weight and dieting, it is not clear what 'thinness' means to anorectics themselves or that norms about dieting are always involved in subjective experiences of anorexia. Meaning-centered studies of anorectics--especially those in non-clinical settings--are needed to clarify the cultural contexts of the disorder. Case studies of two anorectic women from Minneapolis-Saint Paul, Minnesota, show that for some anorectics self-starvation is encoded in religious idioms and symbols about the body, food, and self. A review of the literature illustrates a long-standing relation between self-starvation and religious ideals in Western culture and points to an association between contemporary anorexia nervosa and asceticism. The case studies presented here demonstrate that this asceticism may be subjectively expressed through religious concepts about the body and food and suggest that future research formally investigate the religious practices and beliefs of anorectics seen clinically. The author explores the implications of these findings for definitions of 'normality' and 'abnormality,' key issues in ethnopsychiatry. These findings also suggest that future cross-cultural research might examine asceticism about the body and food in religions other than Judeo-Christian, cultural groups with rituals of fasting and vomiting, and the presence of fundamentalist churches and missionaries in those non-Western cultures for which there are recent reports of eating disorders. Anorexia nervosa's designation as a syndrome limited to Western cultures or to those cultures influenced by them may reflect unexamined assumptions on the part of researchers that dieting and secular ideals of slimness are primarily involved in the disorder.

Adolescent↗
PubMed1982

Amphibian organ culture in experimental toxicology: the effects of paracetamol and phenacetin on cultured tissues from urodele and anuran amphibians.

Organ cultures of various tissues from urodele amphibians deacetylate paracetamol to p-aminophenol, which polymerises to form a brown precipitate. Paracetamol addition results in a loss of glycogen and lactate dehydrogenase (LDH) from urodele liver cultures and an increase in glucose release, and in LDH loss from kidney cultures. Organ cultures from anuran amphibians are unable to metabolise paracetamol and are not affected by its presence in the culture medium. The addition of unpolymerised p-aminophenol resulted in a loss of LDH from urodele and anuran organ cultures, whilst the addition of polymerised p-aminophenol had no such effects. This suggests that the toxic effects which follow the addition of paracetamol to urodele organ cultures are caused by unpolymerised p-aminophenol, a known toxicant in mammals. Cultures from both urodele and anuran amphibians are able to deacetylate phenacetin to p-phenetidine, but p-phenetidine was found to be much less toxic to amphibian tissues than p-aminophenol, causing LDH loss from kidney cultures only at very high dose levels.

Acetaminophen↗
PubMed2005

Cytological and ultrastructural changes induced in anther and isolated-microspore cultures in barley: Fe deposits in isolated-microspore cultures.

To gain further insight into the role played by sporophytic anther tissues in the early stages of the androgenic process, we have compared the cytology and ultrastructure of barley embryogenic pollen grains obtained by anther culture with those obtained by isolated-microspore culture. The microspores behaved similarly in both culture systems but ultrastructural studies detected a significant difference: the presence of electron-dense deposits on the intine of embryogenic pollen grains generated by isolated-microspore culture compared to their absence in grains generated by anther culture. To discover the nature of these deposits, we applied proteinase K and EDTA treatments to ultrathin sections. We also subjected the deposits to X-ray microanalysis and found that they contained iron. Anthers and isolated microspores were cultured in media containing different concentrations of iron so as to evaluate the presence of these deposits on the intine. Deposits were not found in anther cultures at any iron concentration used or in microspore cultures when concentrations were lower than 40 mg/L. The Fe deposits on the intine appear to derive from an excess of Fe in the isolated-microspore culture medium which, if allowed to pass through the cell wall, could well be toxic to the embryogenic development of the microspores.

Cells, Cultured↗