Interline Differences Research Articles

The Derivation and Comparative Characterization of Mesenchymal Stem Cell Lines, Isolated from Human Pulp of a Deciduous Tooth of Children of Different Sexes

Two new non-immortalized fibroblast-like cell lines isolated from the pulp of Deciduous Tooth of Children of Different Sexes, named MSC-DP-1 and MSC-DP-2, were derived and characterized. In order to confirm the status of mesenchymal stem cells, a comparative analysis of a some characteristics in these lines was carried out at early and late passages. In the process of long-term cultivation, significant interline differences were revealed in the nature of replicative senescence (RS) and in growth characteristics. The MSC-DP-1 line was characterized by a later entry into the active stage of RS and more active proliferation compared to the MSC-DP-2 line. Karyotypic analysis showed that both lines have a normal human diploid karyotype at early passages. At the late (18th passage), in the stage of active PS, the MSC-DP-2 line also has a normal karyotype. And the MSC-DP-1 line, which enters the active stage of RS much later (at passage 42), has an abnormal karyotype with a large number of clonal chromosomal rearrangements. Both lines showed a high proportion of cells carrying antigens characteristic of human MSCs: CD44, CD73, CD90, CD105, HLA-ABC, and a low frequency of cells with CD34, CD45, and HLA-DR antigens. The cells of the derived lines at an early passage have the ability to differentiate in adipogenic, osteogenic and chondrogenic directions. But the MSC-DP-2 line shows weaker differentiation in the adipogenic direction, than the MSC-DP-1 line. In the process of RS, a significant weakening of adipogenic differentiation takes place in the MSC-DP-1 line, and it disappears in the MSC-DP-2 line. The activity of other differentiations does not change during RS. In general, the derived results confirm the status of MSCs for the derived lines and indicate interline differences in the RS process. However, the derived differences between these lines, as well as the comparison with the previously derived line MSC-DP do not indicate their gender nature. Apparently, they are associated with the genetic characteristics of different donors.

PO-01-157 MULTIELECTRODE ARRAY RECORDINGS OF IPSC CARDIOMYOCYTES SHOW GREATER VARIABILITY BETWEEN REPLICATE DIFFERENTIATIONS THAN BETWEEN DIFFERENT IPSC LINES FROM HEALTHY INDIVIDUALS AND CORRELATE WITH EXPRESSION OF SARCOMERIC GENES

Induced pluripotent stem cell (iPSC) derived cardiomyocytes are an important model system for cardiac electrical disorders. They offer the potential to detect disease-specific differences in cell lines as well as between healthy individuals of differing genetic backgrounds. However, variability of iPSC cardiomyocyte differentiations makes these studies prone to type 1 errors and optimal experimental designs and sample sizes remain undefined. We aimed to characterise the variability in electrical properties of cardiac myocytes derived from three iPSC lines obtained from healthy individuals, at baseline, in response to varying pacing rates and in the presence of two pharmacological challenges to repolarisation reserve: inhibition of IKr and augmentation of INaL. We aimed to explore the mechanisms underlying the variability in iPSC-derived cardiomyocyte phenotypes by analysing transcriptional profiles. Four replicate cardiomyocyte differentiations from three iPSC lines were analysed on 48-well multielectrode arrays (MEA). Field potential recordings were made from spontaneous beating monolayers as well as during pacing at 0.5, 1.0 and 1.5 Hz at baseline and in the presence of dofetilide (10 nM) and ATX II (25 nM). Field potential parameters were correlated with mRNA levels for a panel of 48 transcripts of key cardiac ion channel, calcium handling, sarcomeric and metabolic genes, assayed using the NanoStrings platform. Significant variability in baseline field potential parameters was observed between replicate differentiations of each line, whereas differences between iPSC lines were not statistically significant in nested ANOVA and mixed effects model analyses after taking into account within line variability. The rate-dependence of repolarisation and repolarisation reserve also showed significant intra-line differences but again interline differences were not significant. Field potential durations showed the strongest correlation with expression of two sarcomeric genes, MYH2 and MYH7. Significant variability in phenotypes between replicate differentiations of the same iPSC line indicates that a large number of replicates are needed in studies designed to detect differences between lines derived from healthy individuals. One should also take this into account when comparing diseased lines with isogenic controls. Maturation state and cardiomyocyte purity of differentiations are an important cause of variability in MEA field potential recordings.

ОПЫТ СЕЛЕКЦИОННО-ПЛЕМЕННОЙ РАБОТЫ С КРАСНО-ПЕСТРОЙ ПОРОДОЙ СКОТА

The article presents assessment results of red-and-white breed lines on milk productivity, live weight and intensity of milk yield. It was found that almost all the descendants of the bulls belonging to the lines of V.B. Ideal 1014315, M. Chieftain 95679, R. Sovering 198998, R. Citation 267150 are characterized by high milk yield. The only exception was the daughters of the bulls from R. Sovering 198998 line, who were inferior to their peers in milk yield by 225-250 kg (4.1-4.5%) during 305 - day lactation. There were no significant interline differences in the content of fat and protein in milk. It was found that positive results were obtained from crosses of V.B. Ideal × M. Chieftain (+204 kg of milk), M. Chieftain × V.B. Ideal (+300 kg), M. Chieftain × R. Sovering (+185 kg), R. Sovering × R. Sheilimar. Intraline selection on milk yield in all cases was ineffective. Assessment of servicing bulls of V.B. Ideal line showed that the improvers of the daughters’ milk yield are the bulls Kumir 1242 (+283 kg to their peers), Leonardo 218 (+244 kg), Jul 43 (+143 kg). Bulls Clemens 12700 (- 201 kg) and Furgon 1268 (- 679 kg) were recognized as deteriorating the milk yield. As for R. Sovering line, the bulls Service 101 (+403 kg) and Mills 284 (+263 kg) are the milk yield improvers, and the bull Berkut 3473 (+0.38%) improves fat content of milk of the daughters.

Effect of long-term emotional-painful stress on the leukocyte composition of blood in rats with different levels of excitability of the nervous system

Post-stress states in animals and humans are accompanied by the development of neuro- and peripheral inflammation. The mechanisms of such immune dysfunctions, their contribution to the pathogenesis of stress-related diseases, as well as the dependence of the intensity of poststress inflammation on genetically determined features of the nervous system, have not been clarified.
 Aim: to assess the dynamics of the development of poststress inflammation depending on the genetically determined level of excitability of the nervous system in rats.
 Materials and methods. The study was carried out on male rats of two lines, selected by the threshold of excitability of the nervous system-the line HT (high threshold of excitability) and LT (low threshold of excitability). As a model of chronic stress, the Protocol of long-term emotional and pain exposure according to Hecht was used. To investigate the dynamics of changes in the leukocyte formula, three time points were selected: 24 hours, 7 days and 24 days after the stressor. Morphological analysis of blood was carried out to determine the leukogram, for which the leukocytes were counted in a blood smear stained by Romanovsky-Gimza.
 Results. Chronic stress leads to an increase in the leukocyte shift index only in the experimental group of highly excitable rats of the LT line compared with control animals of the same line. The significance of the differences is confirmed on day 7 after the end of the stressor. No interline differences in neutrophil/lymphocyte ratios were found in intact animals of LT and HT lines.
 Conclusion. In rats with a genetically determined high level of excitability of the nervous system, post-stressor systemic inflammation appears 7 days after the end of the stressor. Animals with a low level of excitability of the nervous system had no signs of post-stress inflammation throughout the observations. The article discusses the possible mechanisms of detected immune dysfunctions in animals due to high excitability of the central nervous system.

Isolation and Comparative Characteristics of Mesenchymal Stem-Cell Lines Derived from Foreskin of Two Donors of Similar Age

Two new nonimmortalized human cell lines FRSN-1 and FRSN-2 were established from foreskin of two similarly aged donors (2.5 years). Growth characteristics and differentiation potential of these cell lines studied on the sixth passage confirmed their status as mesenchymal stem cells (MSCs). A number of characteristics have been analyzed during long-term cultivation up to the 26th passage. The dynamics of the process of replicative senescence defined by the activity of β-galactosidase differed between these lines. However, at the 26th passage, the process of replicative senescence was equally enhanced in both lines. The plating efficiency markedly differed between the lines on the sixth passage. In FRSN-1, it was higher than in FRSN-2. The plating efficiency substantially dropped to the 26th passage in FRSN-1 and was lost in FRSN-2 line. Growth curves showed active proliferation of these lines at the 6th passage. The average doubling time did not differ between the lines and was 36.9 and 39.0 h, respectively. Analysis of growth curves on the 26th passage revealed a decline in proliferative activity and increase in average doubling time of cell populations in both lines, more in FRSN-2 than in FRSN-1 lines. The patterns of growth curves differed in these lines. Morphological analysis revealed increased cell size and spreading typical for the phase of the replicative senescence. Numerical and structural karyotypic analysis at the sixth passage showed that both lines have normal karyotype 46, XY. We did not discover interline differences in the frequency of chromosomal aberrations. To determine the status of these cell lines, comparative analysis of the surface markers was performed using flow cytometry. It was revealed that cells of both lines expressed surface antigens characteristic of human MSCs: CD44, CD73, CD90, CD105, and HLA-ABC and did not express CD34, CD45 and HLADR. Cells of both lines displayed SSEA-4 and SOX2, markers of human embryonic stem cells (ESCs). Expression of SSEA-4 was also detected at the 26th passage in both lines. FRSN-1 and FRSN-2 cells expressed the markers of early ESC differentiation into three germ layers. The ability of these cell lines to differentiate into osteogenic, chondrogenic, and adipogenic lineages was shown on the sixth passage. Both lines exhibited substantially reduced adipogenic potential on the 20th passage. These data indicate that in contrast to growth characteristics the adipogenic differentiation potential changes even with an average degree of replicative senescence. It appears that the cell replicative senescence contributed to the change in MSC differentiation potential. Overall, the results demonstrate that cell lines derived from different donors are distinguished in growth characteristics and pattern of replicative senescence. The disparity is due to a direct genetic influence and indirectly by different microenvironment in their donor organisms before cell isolation.

Maternally derived egg hormones, antibodies and antimicrobial proteins: common and different pathways of maternal effects in Japanese quail.

Avian eggs contain a variety of maternally-derived substances that can influence the development and performance of offspring. The levels of these egg compounds vary in relation to environmental and genetic factors, but little is known about whether there are correlative links between maternal substances in the egg underlying common and different pathways of maternal effects. In the present study, we investigated genetically determined variability and mutually adjusted deposition of sex hormones (testosterone-T, androstenedione-A4 and progesterone-P4), antibodies (IgY) and antimicrobial proteins (lysozyme) in eggs of Japanese quail (Coturnix japonica). We used different genetic lines that were independently selected for yolk T concentrations, duration of tonic immobility and social reinstatement behaviour, since both selections for behavioural traits (fearfulness and social motivation, respectively) produced considerable correlative responses in yolk androgen levels. A higher selection potential was found for increased rather than decreased yolk T concentrations, suggesting that there is a physiological minimum in egg T levels. Line differences in yolk IgY concentrations were manifested within each selection experiment, but no consistent inter-line pattern between yolk IgY and T was revealed. On the other hand, a consistent inverse inter-line pattern was recorded between yolk IgY and P4 in both selections for behavioural traits. In addition, selections for contrasting fearfulness and social motivation were associated with changes in albumen lysozyme concentrations and an inverse inter-line pattern between the deposition of yolk IgY and albumen lysozyme was found in lines selected for the level of social motivation. Thus, our results demonstrate genetically-driven changes in deposition of yolk T, P4, antibodies and albumen lysozyme in the egg. This genetic variability can partially explain mutually adjusted maternal deposition of sex hormones and immune-competent molecules but the inconsistent pattern of inter-line differences across all selections indicates that there are other underlying mechanisms, which require further studies.

MORPHOLOGICAL CHANGES IN THE HIPPOCAMPUS OF RATS IN ACCELERATED AGING

The aim of this work was the analysis of structural changes with age in the hippocampus of senescenceaccelerated OXYS rats when signs of accelerated brain aging are missing (age 14 days), developments (age 5 months), and active progresses (age 15 months). The study was performed on 15 OXYS rats and 15 Wistar rats (as a control). After dislocation, brains were dissected, fixed with 10% formalin, embedded in paraffin, and serially cut in coronal sections (5μm thickness). These sections were stained with Cresyl violet and examined with a photomicroscope (Carl Zeiss Axiostar plus, Germany). The total number of hippocampal pyramidal cells in the CA1, CA3 and the dentate gyrus regions were estimated in 14-dayold, 5and 15-month-old OXYS and Wistar rats (n = 5) on the 5 slices of each brain sections. The number of neurons with chromatolysis, hyperchromatic with darkly stained cytoplasm and shrunken neurons were calculated as degenerative neurons. The pictures obtained with the program Carl Zeiss Axio Vision 8.0 with increasing 10  100, determined the average area bodies and nuclei of neurons (mkm2). The significant structural changes of neurons in the CA1, CA3 and dentate gyrus regions of the hippocampus in OXYS rats at 5 month of age are revealed by light microscopy. This results indicates the early develop neurodegeneration in OXYS rats. The most pronounced morphological changes occur in the CA1 region of the hippocampus of OXYS rats and irreversible. The degenerative changes of neurons in the hippocampus increases by the age of 15 months. Morphometric analysis of the average area of bodies and the nuclei of hippocampal neurons in CA1, CA3 and the dentate gyrus regions of OXYS and Wistar rats at 14 days of age showed no significant interline differences. At 5 months of age in the CA1 region of the hippocampus of OXYS rats was determined a significantly lower average body size and nuclei of pyramidal neurons compared with Wistar rats. With age, these changes have progressed and 15-month-old OXYS rats have a significant decrease in the area bodies and nuclei in all studied regions of the hippocampus compared with Wistar rats. These results confirm the promising use OXYS rats as a model of accelerated brain.

Differences in the parameters of longevity and its sex-specificity in human populations and modeling them in drosophila

The longevity-related and sex-specific interpopulation variability, as a universal biological phenomenon, occupies a special place among a diversity of unclear phenomena in the biology and genetics of aging and biodemography. The authors carried out a population-genetic variability analysis of some life expectancy (LE) parameters, using a Drosophila model to study the dynamics, sex-specificity of aging, and mortality rates, depending on the geographical origin of Drosophila populations. Changes in the indicated parameters were also studied under stress conditions. Six wild insect lines were studied at the optimal temperature and then separated into three groups according to the life span criterion and the sexual dimorphism differentiation. It is obvious that the initial natural populations were polymorphic in the genes regulating a general longevity level and its sex-specific variability. We detected similar regularities in two laboratory long-inbred standard lines Canton S and Oregon R. The exposure of the initial lines to an elevated and stress-inducing temperature of 29°C shortened approximately by two times the insects’ life span and reduced its interline differences. The lifespan rankings of the lines remained practically the same. However, the nature of sexual dimorphism significantly changed. The longevity-related genes are, probably, environmentally-dependent and most active at 25°C, but their activity is suppressible at stress-inducing temperatures. A comparison of the model and human data allowed us to predict a potential genetic-demographic longevity spectrum and a degree of their stress-induced dependence.

Comparison of blastoderm traits from 2 lines of broilers before and after egg storage and incubation

We examined the impact of egg storage on the hatchability of eggs from 2 lines (A and B) of commercial broiler breeders with known differences in fertility (line B slightly higher) and hatchability following egg storage (line A slightly lower). Eggs from both lines were stored in a 16°C room for 3 to 4, 10 to 12, and 17 d, the blastoderms were isolated, evaluated, and statistically compared. No significant interactions (line × duration of storage) were observed in the following traits: blastoderm diameter, total and percentage of viable blastodermal cells, stage of blastoderm development, and embryo weight and stage of development after 7 d of incubation. However, significant differences were observed when comparing line averages across storage treatments (diameter and total number of blastodermal cells) and storage treatment averages across lines (percentage of viable cells, and embryo weight and stages following incubation). To ascertain the basis for the observed differences in fertility, perivitelline sperm-hole numbers were determined in eggs within 72 h of lay (3 to 4 d storage group). Eggs from line B had a significantly higher number of eggs having greater numbers of sperm holes than eggs from line A. It is concluded that no single blastoderm trait or combination of traits can be definitively associated with the inter-line differences in hatchability following storage of eggs. Alternatively, differences in PVL sperm-hole counts between lines may be associated with the reported differences in fertility between the 2 lines.

Maximal inflammatory response benefits syngeneic skin graft acceptance

We investigated the influence of acute inflammation in skin isograft acceptance. Two mouse lines selected for maximal (AIRMAX) or minimal inflammatory response (AIRMIN) were transplanted with syngeneic skin. Cellular infiltrates and cytokine production were measured 1, 3, 7 or 14 days post-transplantation. The percentage of CD4+CD25+Foxp3+ cells in the lymph nodes was also evaluated. Grafts were totally accepted in 100% of AIRMAX and in 26% of AIRMIN mice. In the latter, partial acceptance was observed in 74% of the animals. Emigrated cells were basically PMN and were enhanced in AIRMAX transplants. IL-10 production by graft infiltrating cells showed no interline differences. IFN-gamma was increased in AIRMIN grafts at day 14 and lower percentages of CD4+CD25+Foxp3+ cells in the lymph nodes were observed in these mice. Our data suggest that differences in graft acceptance might be due to a lack of appropriate regulation of the inflammatory response in AIRMIN mice compromising the self/non-self recognition.

Divergent selection for shape of growth curve in Japanese quail. 4. Carcase composition and thyroid hormones

1. Changes in the relative weights of carcase, abdominal fat, breast and leg muscles, and plasma thyroid hormone concentrations occurring during the first 6 weeks of postnatal growth were analysed in males of HG and LG lines divergently selected for high and low relative body weight (BW) gain between 11 and 28 d of age, respectively, and constant adult BW. 2. The second week of postnatal life was a critical age at which the HG males exhibited a relatively faster growth in comparison to their LG counterparts and permanently exceeded LG males in the percentage by weight of carcase, breast and leg muscle. A higher production of muscle tissues was associated with lower accumulation of abdominal fat before sexual maturity. 3. In general, the plasma T3 level of HG quail exceeded that of LG quail. Nevertheless, significant differences were found only at 14, 21 and 28 d of age, that is, in the period during which the highest inter-line differences in relative growth rate were noted. Also the T3/T4 ratio followed a similar trend while plasma T4 level showed no clear and consistent inter-line differences. 4. The results suggest that the selection for the shape of the growth curve, like the selection for body fat, modifies the carcase quality owing to shortening/prolongation of the acceleration growth phase. Individuals with a short acceleration phase of the growth curve are characterised by low carcase quality during the fattening period.

Lack of correlation between rabies virus replication in the brain and antibody isotype profile in genetically modified mice

The relationship among the phenotypes resistance to infection, virus replication in the brain and isotype production was investigated in genetically modified High (H) or Low (L) antibody responder mouse lines. Although they express the same innate susceptibility to rabies infection, these lines differ as to different viral replication rates in the central nervous system and L mice showed a higher permissible state. After intramuscular infection with the Pasteur rabies strain (PV), the H-L interline differences on the earlier stage of virus replication were 1000 and 80 folds on days 5 and 6, respectively. The isotype profile in sera of the experimentally infected mice reflected an interline difference of 25 folds for IgG2a throughout the infection period, and for the IgE production the H-L difference was highly significant only at the beginning of the process. These results confirm the multi-specific effect of antibody immune responsiveness and the general isotype distribution of antibodies in these genetically selected mice. Contrary to the clear correlation between antibody responsiveness and the acquired resistance to rabies infection, the present study demonstrates that the constitutive genetic character of High and Low responder individuals does not intervene in the degree of resistance following infection. Altogether, this study contributes to the knowledge of the protective role of the general innate responsiveness on the pathological pattern to rabies virus infection.

Immunological, Cytogenetical, and Behavioral Changes in CBA and C57BL/6 Male Mice after Pheromonal Action

Immunological and cytogenetical changes were studied in mice-recipients of CBA and C57BL/6 lines as well as modifications of their behavior appearing as a result of action of various stressors. Post-stress volatile excretions of CBA males cause led to a pronounced immunosuppression in males-recipients of both lines. This resulted in a decrease of capability for immune response of the stressed mice to sheep erythrocytes. Besides, in young males of the CBA line, frequency of chromosome aberrations in bone marrow cells increased statistically significantly after stressing by syngenec volatile excretions of mature animals. Attractive properties of volatile excretions of the CBA and C57BL/6 line males were studied. In norm, animals of both lines prefer syngenic odors of intact males donors. However, for the CBA line males, post-stress excretions from males-donors of both lines became more attractive than excretions from intact animals. Also revealed were interline differences in incidence of choice of syngenic and allogenic post-stress excretions. This can be explained both by different stress-reactivity of animals-donors and differences of recipients of the studied genotypes in selectivity to olfactory stimuli. An importance of olfactory stresses and of the revealed interline differences for explanation of some micro-and macroevolutionary synecological processes in the domestic mouse is discussed.

Cancer professionnel : intérêt d’une consultation de cancérologie du travail

Wistar derived inbred line, the WAG/Rij rats, genetically absence epilepsy prone and their normal counterparts, outbred Wistar rats, were compared in respect to differences in behavior, in acute and chronic antidepressant imipramine treatment and in the immediate early gene c-fos expression in the brain regions induced by forced swimming test procedure. The WAG/Rij rats as compared with Wistar rats were found to exhibit decreased activity in the open field test, increased immobility in the forced swimming test and decreased sucrose intake (anhedonia). Interline differences indicating increased anxiety in the WAG/Rij rats were not revealed in the light–dark choice, social interaction and elevated plus-maze tests. The WAG/Rij rats in contrast to Wistar rats responded only to chronic antidepressant imipramine treatment with a reduction in their enhanced immobility in the forced swimming test. “Behavioral despair” induced by forced swimming led to c-fos expression in frontal cortex, nucleus accumbens and striatum, terminal regions of three dopaminergic brain systems (mesocortical, mesolimbic, nigrostriatal). The c-fos expression in the brain of WAG/Rij rats was substantially higher than that of Wistar rats. Moreover, the strains differed in the distribution of c-fos expression between brain regions. Results suggest that WAG/Rij rats are prone to adopt passive strategies of behavior in stressful situations, and so in this certain aspect this strain might be regarded as new experimental (genetic) model of depressive-like (passive) behavior accompanying absence epilepsy. Further testing this hypothesis is proceeding. This putative model could be used for the investigation of neurobiological basis and mechanisms of such “double pathology” and for the examination of new concepts of its therapy.

Effects of Lonomia obliqua (lepidoptera, saturniidae) toxin on clotting, inflammatory and antibody responsiveness in genetically selected lines of mice

Lines of mice genetically selected for high (H) or low (L) antibody response and for maximal (AIR MAX) or minimal (AIR MIN) acute inflammatory reaction, in which the opposite extreme potentialities have been clearly defined, offer an appropriate model for investigating the environmental and genetic factors acting on innate and adaptative immunobiological functions. This model has been successfully employed to study the resistance or susceptibility against pathogens and/or toxins. It had been demonstrated that the skin contact with Lonomia obliqua caterpillar bristles induces local inflammation and may elicit severe hemorrhagic disorders. In the present study, blood coagulation time, and the acute inflammatory reaction were scored 24 h after injection of the Lonomia bristles crude extract in a subcutaneous dorsal air pouch. The acute inflammation was determined by the leukocyte concentration in the local exudates. The highest interline differences were observed between the AIR MAX (10 6 cells/ml) and AIR MIN (2×10 5 cells/ml) and this distinct expression involves the number of monocytes, eosinophils and mainly neutrophils. Regarding coagulation, the highest interline difference was observed between the H III and L III mice, and the F 1 [L III×H III] hybrids showed the overdominance of the fast clotting character. The adaptative immune response was evaluated by comparing the anti- Lonomia bristle extract IgG titer among the lines: the antibody titers were higher in the H lines than in the L ones and equivalent in the AIR MAX and AIR MIN mice, in accordance to the phenotype profiles generated by the distinct selective processes. The genetically selected mice lines—AIR MAX, AIR MIN, H I, H III, H G, L III and L G—showed an almost continuous distributions for inflammation, coagulation time and IgG antibody titers, being the interline variances always higher than the intraline ones for the individually measured phenotypes. Altogether, these results suggest the independent polygenic regulation of these traits, being indicative of the genetic control to Lonomia toxin innate and adaptative sensitivity in humans.

Divergent selection for shape of growth curve in Japanese quail. 2. Embryonic development and growth

1. Embryonic growth and development were analysed using meat type lines of Japanese quail, HG and LG, divergently selected for shape of the growth curve. A total of 1020 embryos of generations 9, 10 or 13 were used for analysis. 2. Considerable inter-line differences were observed in the rate of embryonic development. When compared to HG, LG embryos appeared to be developmentally accelerated during the first 42 h of incubation (larger blastoderm diameter, more somites and higher frequency of more advanced Hamburger–Hamilton stages) as well as at the end of the prenatal period (more embryos with the yolk sac inside the body cavity, shorter incubation period). This corresponded with the trend in postnatal development. 3. Embryonic growth of both lines exhibited an exponential trend. However, considerable inter-line differences were noted in the rate of embryonic growth. Initial growth retardation compensated subsequently by a higher growth rate of HG vs LG quail, characterised the lines after hatching. The same growth pattern repeated three times during the prenatal period (between d 0 and 3, 3 and 8, and 8 and 16). 4. The repeated occurrence of transient decreases in growth rate of the developmentally delayed HG line could be associated with a delayed onset of genetically determined physiological functions mediating utilisation of nutrient supply. 5. Hence, different shapes of growth curves in two genotypes with similar growth potential reveal inter-line differences in physiological age persisting during the whole ontogenesis.

The process of embryonic mortality in seven initial laying strains during incubation

Abstract. In five segments of incubation (within the 4th day, on the 7th, 14th, 20th days and in the stage of breaking through the shell) we studied the fertility of hatching eggs, the hatchability and embryonic mortality of seven initial laying strains (RIR – 1, RIR – 2, RIR – 3, RIW – 1, SU – 1, BPR – 1 and BPR – 2) in the course of three egg sets of individual pedigree hatching. The average hatchability of all the seven strains was 71.5%, the proportion of unfertilised eggs was 11.3% and embryonic mortality reached 17.2%. The embryonic mortality of all the seven strains culminated in two periods, i.e. within the 4th day of incubation (36.9% of the total embryonic mortality) and at the end of incubation (from the 15th to 20th day of incubation – 23.4% and during breaking through the shell – 18.1%). In terms of the inter-line differences, the embryonic mortality was the highest in three lines, which had a higher coefficient of inbreeding (Fx < 12.5%), i.e. RIR – 1 (20.9%), BPR – 2 (20.3%) and RIW – 1 (23.7%). All the three lines bear the recessive gene (k+) on locus K, which is responsible for the rapid type of feathering using feathersexing. Within the 7th day of incubation the embryonic mortality was higher in both BPR lines (BPR – 1–58.1%, BPR – 2–53.6% of the total embryonic mortality), while in the last trimester of incubation and in the period of breaking through the shell it was the lowest (BPR – 1–30.8%, BPR – 2–36.2% of the total embryonic mortality).

Depressive-like behavioral alterations and c-fos expression in the dopaminergic brain regions in WAG/Rij rats with genetic absence epilepsy

Wistar derived inbred line, the WAG/Rij rats, genetically absence epilepsy prone and their normal counterparts, outbred Wistar rats, were compared in respect to differences in behavior, in acute and chronic antidepressant imipramine treatment and in the immediate early gene c-fos expression in the brain regions induced by forced swimming test procedure. The WAG/Rij rats as compared with Wistar rats were found to exhibit decreased activity in the open field test, increased immobility in the forced swimming test and decreased sucrose intake (anhedonia). Interline differences indicating increased anxiety in the WAG/Rij rats were not revealed in the light–dark choice, social interaction and elevated plus-maze tests. The WAG/Rij rats in contrast to Wistar rats responded only to chronic antidepressant imipramine treatment with a reduction in their enhanced immobility in the forced swimming test. “Behavioral despair” induced by forced swimming led to c-fos expression in frontal cortex, nucleus accumbens and striatum, terminal regions of three dopaminergic brain systems (mesocortical, mesolimbic, nigrostriatal). The c-fos expression in the brain of WAG/Rij rats was substantially higher than that of Wistar rats. Moreover, the strains differed in the distribution of c-fos expression between brain regions. Results suggest that WAG/Rij rats are prone to adopt passive strategies of behavior in stressful situations, and so in this certain aspect this strain might be regarded as new experimental (genetic) model of depressive-like (passive) behavior accompanying absence epilepsy. Further testing this hypothesis is proceeding. This putative model could be used for the investigation of neurobiological basis and mechanisms of such “double pathology” and for the examination of new concepts of its therapy.

Co-localization of quantitative trait loci regulating resistance to Salmonella typhimurium infection and specific antibody production phenotypes

Salmonella enterica serotype typhimurium is a facultative intracellular bacteria that induces systemic infection in mice. Resistance to this pathogen is under polygenic control in which Nramp1 is the major gene involved. Lines of mice obtained by selective breeding for high (HIII) or low (LIII) antibody response to flagellar antigens of salmonellae showed significant susceptibility differences, although both the lines display Nramp1 R alleles. The HIII line was extremely susceptible to infection, while the LIII line was resistant. In order to examine the cellular and genetic mechanisms involved in this distinct pattern of resistance, HIII and LIII mice were analyzed for IFNγ and IL4 production and screened for quantitative trait loci involved in S. typhimurium infection, using several polymorphic microsatellites. In the present work, HIII mice showed an IFNγ downregulation in the early phase of infection when compared with LIII animals. No interline differences in IL4 production were verified. The loci screening was performed on immunized F2 intercrosses obtained from HIII and LIII mice. Three antibody-controlling chromosomal regions were coincident, and another was mapped near one of the four loci known to affect susceptibility to S. typhimurium. These results indicate a major role of IFNγ in our model, and suggest the co-localization of quantitative trait loci modulating both infection and antibody production phenotypes.

Comprehensive galectin fingerprinting in a panel of 61 human tumor cell lines by RT-PCR and its implications for diagnostic and therapeutic procedures.

Knowledge about galectin expression by human tumor cells is mainly restricted to galectins-1 and -3. This study was conducted to define the gene expression pattern of all presently known human galectins in tumor cell lines of various histogenetic origin (galectinomics). The presence of mRNAs for human galectins-1, -2, -3, -4, -7, -8, and -9 was monitored by RT-PCR analyses in a panel of 61 human tumor cell lines of different origin (breast, colon, lung, brain, skin, kidney, urogenital system, hematopoietic system). The validity of the technique was first confirmed by comparison of RT-PCR data with those obtained by Western blotting and cytofluorometry for galectins-1 and -3 in 18 cell lines. The following detection of a complex pattern of gene expression beyond commonly studied galectins-1 and -3 underscored the need for this fingerprinting. The most abundantly expressed message for a member of this lectin family was galectin-8 with 59 positive cell lines. With the exception of the tested lung tumors, galectin-1 and -3 transcripts were frequently expressed in the cell line panel with differences between individual cases. Positivity for galectins-2 and -4 was confined to a significant fraction of colorectal and neural tumors. Signals for galectin-9, the third known human tandem-repeat-type galectin besides -4 and -8, appeared in colorectal carcinoma cell lines with a frequency similar to that of galectin-4 but with inter-line differences. Its expression was restricted to lines of this tumor type, of the tested ovarian carcinoma, and hematopoietic malignancies. The results clearly demonstrate that human tumor cells express more mRNA species for galectins than those for galectins-1 and -3. To derive unequivocal diagnostic and prognostic information by immunohistochemistry on galectins with antagonistic impact on growth control and significant influence on cell adhesion, additional monitoring of these so far insufficiently studied family members is essential.