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PubMed1996

Altered growth and attachment of rabbit crypt colonocytes isolated from different developmental stages.

The rabbit colon was used to establish an in vitro model for examining development-related cellular changes in colonocyte function. Colonic epithelia from newborn, weanling, and adult animals were separated from the muscle and subjected to enzymatic digestion. A mixture of 0.05% Pronase, 0.015% collagenase IV, and 0.023% DTT was determined to be optimal for the isolation of newborn and weanling colonocytes. This solution yielded significantly more cells and of greater viability than a 0.1% Pronase, 0.03% collagenase IV, 0.07% DTT mixture that is optimal for adult colonocytes. The epithelial origin of the colonocytes was confirmed by immunofluorescent staining of cytokeratins. The isolation procedure resulted in a crypt-enriched population and the cell yield/g of mucosa increased with age as did the crypt depth. Colonocyte viability of adults but not of newborns and weanlings, declined from 24 to 72 h. When grown on plastic, the newborn and weanling colonocytes show a approximately 2-fold increase in number, DNA and protein content over 48 h. In contrast, for all three parameters the adult colonocytes revealed only a approximately 10% increase. The colonocytes also showed an age-related decline in attachment to extracellular matrices. Colonocytes showed maximal attachment to Matrigel and collagen IV; newborn and weanling colonocytes show > 80% attachment, whereas adult colonocytes showed only a 45% attachment. The efficacy of attachment to Matrigel compared with that on plastic also differed with age, representing 9.3-, 5.5-, and 4.4-fold increase in adult, weanling, and newborn colonocytes, respectively. Newborn and weanling colonocytes grown on Matrigel for 48 h, showed a significant, 15% increase in cell number, DNA, and protein content compared with those grown on plastic. There was no difference in these parameters when adult colonocytes grown on Matrigel were compared with those grown on plastic. In summary, we have established an in vitro model for studying colonic epithelial cells at different stages of development.

Age Factors↗
PubMed1983

Comparison of cytochrome P-450-dependent metabolism in different developmental stages of Drosophila melanogaster.

The activities of several drug metabolizing enzymes were compared in microsomes from larvae and adult Drosophila. The cytochrome P-450 content and the benzo[a]pyrene (BP) hydroxylation, p-nitroanisole demethylation and 3- and 4-hydroxylation of biphenyl were 4-20-fold higher in microsomes from adult flies, while 7-ethoxycoumarin deethylase activity and cytochrome c reductase activity were about the same in the two stages. 2-OH-biphenyl was formed in trace amounts by microsomes from adult flies but not to any detectable amount by microsomes from larvae. Pretreatment with phenobarbital (PB), Aroclor 1254 (PCB) or beta-naphthoflavone (BNF) increased the cytochrome P-450 content and the various cytochrome P-450-mediated reactions up to 7-fold in larvae. The effects of the pretreatments were weaker in adult flies, where the increase never was more than 3-fold, and many reactions were unaffected by the pretreatments. BNF was thus inefficient in enhancing all reactions, except a slight (1.3-fold) increase in the formation of 4-OH-biphenyl. Microsomes from both stages exhibited increases in specific protein bands with apparent molecular weights of 51 000-58 000 in the sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis following treatment with PB, PCB and BNF. Differences were observed between larvae and adults with respect both to the number of and the molecular weights of the increased protein bands.

Animals↗
PubMed1989

Partial denervation of the rat soleus muscle at two different developmental stages.

Rat soleus muscles were partially denervated at two developmental stages. The L5 ventral ramus was sectioned in rats which were 4-6 days old, when the motor unit size of soleus muscles was still large, and at 17-19 days, when motor unit territory reached its adult value. The response of axons in the L4 ventral ramus to this procedure was then investigated. The removal of the L5 ventral ramus at 4-6 days results in an initial brief increase of motor unit size, after which the motor units retain the territory they occupied at 4-6 days. After removal of the L5 ventral ramus at 17-19 days, the L4 ventral ramus is able to expand to occupy a territory comparable in size to that of animals operated at 4-6 days. In both cases the final percentage of mean motor unit tension is two- to three-fold greater than that in normal muscles. Although the final motor unit territory is similar for both groups, it is achieved by different mechanisms. In animals operated on at 4-6 days the normal elimination of terminals does not occur, and the large neonatal motor units are retained, whereas in animals operated on at 17-19 days the peripheral field of L4 axons expands by axonal sprouting.

Aging↗
PubMed2006

A genome scan for quantitative trait loci associated with body weight at different developmental stages in chickens.

A genome scan to detect quantitative trait loci (QTL) affecting body weight in chickens was conducted on 238 F(2) chickens from a reciprocal cross of Silky Fowl and White Plymouth Rock using 125 microsatellite markers covering 23 autosomes and the Z chromosome. Two types of QTL were considered: static QTL (SQ) and developmental QTL (DQ). Static QTL affected the body weight from hatch to time t, and DQ affected the body weight from time t-1 to time t. Six SQ and nine DQ were detected. Of these QTL, four reached a genome-wide significance of 5% or better, with SQ1 and DQ1 being the most significant QTL. Static QTL1 was on chromosome 1 between GCT0006 and MCW0106 and explained 4.05-9.80% of the phenotypic variation in body weights from 3 to 12 weeks of age. At 9, 10 and 11 weeks, the genome-wide significance thresholds of SQ1 were <1%. Developmental QTL1 was located on chromosome 1 between MCW0168 and GCT0006, and explained 2.75% of the phenotypic variation for body weight from week 7 to 8 with a genome-wide significance level <1%. The results suggest that body weight from hatch to time t and developmental growth from time t-1 to time t may involve two different sets of genes or gene actions.

Animals↗
PubMedDate not supplied

Different effects of neuroprotectants FK-506 and cyclosporin A on susceptibility to pilocarpine-induced seizures in rats with brain injured at different developmental stages.

Susceptibility of the injured brain to epileptic seizures depends on the developmental stage at which the injury had been inflicted (our previous paper published in Epilepsy Res. 53 (2003) 216-224). The present study was designed to examine whether neuroprotective agents applied following the injury can decrease the seizure susceptibility. In order to solve this problem, the left cerebral hemisphere was mechanically injured in 6- and 30-day-old Wistar rats. Neuroprotectants FK506 or Cyclosporin A (CsA) were injected 20 min and 24h following the injury. On postnatal day 60, all the animals received single i.p. pilocarpine injections to evoke epileptic seizures. During a 6h period following the injection, the animals were observed continuously and pilocarpine-induced symptoms were recorded and rated. The animals were sacrificed 7 days after pilocarpine injection. In rats injured on postnatal days 6 or 30 (P6 or P30, respectively) and injected with FK-506 after the injury, signs of amelioration in the course of epilepsy were observed. Generally, proportions of rats suffering from heavy seizures were lower and/or their survival periods were longer. Following treatment with CsA, proportions of rats displaying heavy seizures were greater. It was accompanied by extremely high mortality (in rats injured on P6) or a longer duration of seizures (in rats injured on P30). The results appear to point to age-dependent differences between the mechanisms of action of the two neuroprotectants.

Aging↗
PubMed1996

Apparent apoptotic cell death in the olfactory epithelium of adult rodents: death occurs at different developmental stages.

In the olfactory epithelium of adult rodent, receptor neurons are generated continually. Despite the ongoing generation of new neurons, no corresponding increase occurs in the thickness of the mature olfactory epithelium. Thus, epithelial cell death must occur to offset the continual generation of new cells. In the present study, a sensitive method to label nicked DNA in dying cells was combined with immunocytochemistry to determine the identity of dying olfactory cells. In addition, the positions of putative apoptotic cells were mapped to provide additional information about the identity of dying cells. Double labeling experiments revealed that each of the olfactory cell types, i.e., basal cells (keratin-positive), immature neurons (GAP43-positive) and mature receptor neurons (olfactory marker protein (OMP)-positive), were positive for fragmented DNA, suggesting that they undergo apoptotic cell death. The results of the mapping study suggest that apoptotic cell death occurs primarily among GAP43-positive neurons.

Animals↗
PubMed1986

Proto-oncogene c-myc is expressed in cerebellar neurons at different developmental stages.

During post-natal cerebellar development the steady-state levels of c-myc transcripts exhibit characteristic changes. As determined by the S1 nuclease protection assay the level of c-myc transcript, which is very high in the late embryonic cerebellum, decreased to low levels shortly after birth. One week later there is a second period of c-myc mRNA accumulation followed by a marked decline to finally reach the low adult value. The second peak of high c-myc mRNA level correlates well with the proliferation of granule cell precursors, and it is characterized by a marked change in the ratio of the two types of transcripts started at the known c-myc promoters 1 and 2. This indicates a change in the cell population involved in the transcription of the c-myc gene. In situ hybridization shows transiently elevated c-myc mRNA levels in neurons of the cerebellar cortex. At post-natal days 3 and 10 (P3 and P10) c-myc transcripts are detectable in the superficial external granular layer composed primarily of mitotically active (neural precursor) cells. Purkinje cell somata show cytoplasmic label at P10. These large postmitotic neurons undergo rapid differentiation at this developmental stage. In the adult cerebellum the low c-myc mRNA level is apparently due to Purkinje cells with barely detectable amounts of c-myc transcripts. The vast majority of mature cerebellar neurons, the internal granule cells, have no specific hybridization signal for c-myc. We conclude that neurons in vivo can accumulate c-myc messenger during proliferation and/or differentiation, perhaps as a cellular response to an external signal.

Aging↗
PubMed1990

Precocious induction of tyrosine aminotransferase mRNA by hydrocortisone in cultured fetal rat hepatocytes at different developmental stages.

Tyrosine-aminotransferase (TAT) is encoded by a liver-specific gene known to be expressed perinatally. Fetal rat hepatocytes (gestation day 19) in primary cultures, in which TAT gene expression is normally undetectable, are induced by hydrocortisone to express TAT-mRNA in a dose-dependent manner (greater than 10(-7) M). In hepatocytes incubated with hydrocortisone, TAT-mRNA levels were marginal after 24 hours, reaching maximal levels at 48 hours. After a pre-incubation of hepatocytes for 24 hours in the absence of hydrocortisone followed by exposure to hydrocortisone (24-48 hours). TAT-mRNA levels were high. Hepatocytes derived from fetuses of gestation days 14 and 17 displayed comparable levels of TAT-mRNA in response to hydrocortisone. These results demonstrate that cultured hepatocytes of gestational stages as early as day 14, which initially do not respond to hydrocortisone by TAT gene induction, undergo a "maturation" process during the initial 24 hours following cultivation, resulting in the acquisition of precocious competence for TAT gene transcription in response to hydrocortisone. This suggests that one or more factor(s), required for hydrocortisone-inducible TAT gene transcription, and not available in fetal liver until birth (Gluecksohn-Waelsch: Cell, 18:225-237, 1979) appear in fetal hepatocytes upon cultivation during this "maturation" period, thus permitting precocious TAT gene expression in vitro.

Animals↗
PubMed2003

Comparative analysis of nuclear proteins of B cells in different developmental stages.

The developmental stage-specific regulation of V(D)J recombination, a gene rearrangement process of antigen receptor gene segments, is tightly controlled in cells. Here we screened proteins uniquely or differentially expressed among three developmentally distinguishable B cells (pro-B, pre-B and mature B cells) using two-dimensional gel electrophoresis and mass spectrometry. Chromatin assembly factor 1 was uniquely expressed in pro-B cells. Purine nucleotide phosphorylase, LCK, E2A and many other unidentified proteins were dominantly present in the nucleus at the early stage of B cell development where the V(D)J recombination process occurs. Also, few proteins including guanidine nucleotide binding proteins were exclusively expressed in pre-B cell. Such findings can provide some information to help understand the developmental regulation of gene rearrangement occurring during B cell development.

Animals↗
PubMed1998

Generation, identification, and evaluation of expressed sequence tags from different developmental stages of the Asian blood fluke Schistosoma japonicum.

Here we report 658 expressed sequence tags (ESTs) generated from the 5'-termini of clones randomly selected from directional cDNA libraries constructed from mRNAs from three developmental stages of Schistosoma japonicum. Putative identifications were assigned to 46. 2% of the ESTs; 6.4% were previously known from S. japonicum, 5.6% were previously known from S. mansoni, 34.2% were known from other organisms, and the remaining 53.8% may represent S. japonicum-specific genes. These 658 ESTs appeared to be derived from 457 unique genes, which together represent 2 to 3% of the 15,000 to 20,000 genes predicted to occur in the schistosome genome.

Animals↗
PubMed1997

Modulatory subdomains of the HS2 enhancer differentially regulate enhancer activity in erythroid cells at different developmental stages.

The HS2 enhancer in the locus control region of human beta-like globin genes displays developmental-stage-independent enhancer function. The mechanism by which it regulates the transcription of the globin genes in erythroid cells throughout development is not fully understood. In this paper we dissect the HS2 enhancer into an enhancer core and five modulatory subdomains M1 to M5. The enhancer core possesses developmental-stage-independent enhancer activity. The modulatory subdomains by themselves do not possess such enhancer activity, but they apparently respond to environmental signals and modulate enhancer core activity in a developmental-stage specific manner. M1 located 5' of the core strongly stimulates core activity in K562 cells at the embryonic stage. M2 and M3 located 3' of the core strongly stimulate core activity in MEL cells at the adult stage. Moreover, M3 suppresses core activity at the embryonic stage and exhibits an adult-stage-selector activity. These findings indicate that the apparent developmental-stage-independence of the HS2 enhancer is a result of multiple interactions between the core and the modulatory subdomains located both near and far from the core.

Adult↗
PubMed1993

Osmometric behavior, hydraulic conductivity, and incidence of intracellular ice formation in bovine oocytes at different developmental stages.

Bovine oocytes that were immature (IMM), matured in vitro (IVM) or in vivo (MAT), or matured and fertilized in vitro (IVF) were studied using a microscope diffusion chamber to estimate osmotic parameters and a cryomicroscope to characterize intracellular ice formation (IIF). Linear Boyle van't Hoff relationships were observed with all four types of oocytes between 0.265 and 0.799 osm NaCl. At 20 degrees C, estimates of hydraulic conductivity (Lp) were significantly higher for IVM oocytes than IMM and MAT oocytes (0.84 micron/(min.atm) vs 0.45 and 0.47, respectively). IVM oocytes also tended to have higher Lp values than IVF oocytes (0.55 micron/(min.atm)). At 5 degrees C, the Lp of IVM oocytes decreased to 0.36 micron/min.atm) corresponding to an Arrhenius activation energy of 7.84 kcal/mol. The incidence of IIF in MAT oocytes suspended in salt solution and subjected to linear cooling to -60 degrees C was 45% at 4 degrees C/min, 75% at 8 degrees C/min, and 93% at 16 degrees C/min; with IVF oocytes, the incidence of IIF was 40% at 4 degrees C/min, 92% at 8 degrees C/min, and 100% at 16 degrees C/min. Comparisons involving median IIF temperatures (TIIF50s) and the distributions of the observed IIF temperatures for IMM (Myers et al., Cryo-Lett. 8, 260), IVM (Chandrasekaran et al., Cryobiology 27, 676), MAT and IVF oocytes indicated that the IIF incidence in IMM oocytes cooled at 4 degrees C/min was greater than that of oocytes at the other developmental stages cooled at the same rate. The TIIF50s of IVM and IVF oocytes were lowered by equilibration in 1.5 M ethylene glycol (EG), glycerol, or propylene glycol (PG) prior to cooling, with EG tending to lower the TIIF50s more than glycerol or PG. For all three cryoprotectants, the TIIF50s and IFF temperature distributions were cooling-rate dependent. The Weibull probability distribution was fitted to the distributions of the IIF temperatures of oocytes suspended in salt solutions with and without cryoprotectants yielding R2 values ranging from 0.70 to 0.98.

Animals↗
PubMed1999

Identification of regulatory regions driving the expression of the Drosophila spalt complex at different developmental stages.

The zinc finger transcription factors Spalt and Spalt-related have been implicated in multiple developmental processes. In the wing they are regulated by the secreted protein Decapentaplegic and participate in the positioning of the wing veins. The function of Spalt has been also analyzed during tracheal development and embryonic segmentation. Here, we present the isolation and characterization of novel spalt/spalt-related alleles, which analysis indicates that these genes cannot substitute for each other in the developmental processes studied. The mutants present embryonic or pupal lethality, with phenotypes consistent with the loss of spalt function. We also present a detailed functional analysis of the DNA regions implicated in the regulation of these genes. This regulation is complex, integrating the information from both negative and positive regulators, and it is modular, with discrete fragments of DNA directing expression to discrete regions in embryonic and larval tissues.

Alleles↗
PubMed1998

Cadmium uptake and bioaccumulation in Xenopus laevis embryos at different developmental stages.

The uptake of cadmium in Xenopus laevis embryos was studied by exposing them to solutions containing cadmium concentrations ranging from 0.1 to 2 mg Cd2+/L at seven developmental stages for 72 h. The uptake values were from 0.0027 microgram Cd2+/embryo (two blastomeres stage) to 0.081 microgram Cd2+/embryo, (hindlimb bud distinct stage). During early developmental stages, a limited permeability to cadmium could play a significant role for the survival of embryos. At the last developmental stage evaluated there was a significantly higher uptake of cadmium simultaneously with a very significant increase in the resistance against cadmium toxicity. The cadmium bioaccumulation factor (BF) ranged from 5 to 460. As a general pattern at all developmental stages the BF was higher in embryos exposed to the lower cadmium concentration, and as development advanced the BF increased.

Animals↗
PubMed1991

Detection of different developmental stages of malaria parasites by non-radioactive DNA in situ hybridization.

A highly sensitive non-radioactive DNA in situ hybridization procedure is described that enables detection and unequivocal identification of various developmental stages of human and rodent malaria parasites. Using biotinylated species-specific DNA probes, erythrocytic parasites can be specifically stained in blood smears. Similarly exoerythrocytic stages can be visualized in cell culture and in sections of paraffin-embedded liver. In blood smears, the hybridization procedure provides a rapid detection of (low) parasitemia and species-determination for experienced microscopists at 100 to 400x magnification. Moreover, the procedure can be applied even after previous Giemsa staining of the preparation, enabling revision of patient smears which were difficult to read after routine Giemsa staining.

Animals↗
PubMed1977

Changes in the thermal denaturation profiles of DNA from different developmental stages of the newt Triturus vulgaris.

The melting profiles of DNA samples from the early gastrula and early neurula of Triturus vulgaris are essentially the same, whereas DNA from mid to late gastrula possesses higher Tm values and shows a deviation from the regular sigmoidal shape at temperatures above Tm. The plot on normal probability paper indicates a second DNA fraction which melts at higher temperatures and, consequently, it has a higher GC-content than the bulk of DNA. These facts confirm our idea that differential DNA replication occurs during gastrulation.

Animals↗