Liver regeneration in dogs: Morphologic and chemical changes

Forty-four percent and 72% hepatectomy were carried out in dogs and the animals were sacrificed for biochemical and pathologic studies from 0.5 to 6 days later. Compensatory hypertrophy and hyperplasia ("regeneration") were evident within 1 day, reached a maximum in 3 days, and were almost complete by 6 days. Coincident with the histologic events of regeneration were decreases in responsiveness of receptor adenyl cyclase to glucagon stimulation, increases of cyclic AMP, inconsistent changes in plasma insulin, and increases in plasma glucagon. These studies have standardized hepatic resection in dogs and they have focused attention upon some possible mechanisms that will require further study. © 1978 Academic Press, Inc. All rights of reserved

High-resolution quantitative imaging of mammalian and bacterial cells using stable isotope mass spectrometry

Background: Secondary-ion mass spectrometry (SIMS) is an important tool for investigating isotopic composition in the chemical and materials sciences, but its use in biology has been limited by technical considerations. Multi-isotope imaging mass spectrometry (MIMS), which combines a new generation of SIMS instrument with sophisticated ion optics, labeling with stable isotopes, and quantitative image-analysis software, was developed to study biological materials. Results: The new instrument allows the production of mass images of high lateral resolution (down to 33 nm), as well as the counting or imaging of several isotopes simultaneously. As MIMS can distinguish between ions of very similar mass, such as ^{12}C^{15}N^{-} and ^{13}C^{14}N^{-}, it enables the precise and reproducible measurement of isotope ratios, and thus of the levels of enrichment in specific isotopic labels, within volumes of less than a cubic micrometer. The sensitivity of MIMS is at least 1,000 times that of ^{14}C autoradiography. The depth resolution can be smaller than 1 nm because only a few atomic layers are needed to create an atomic mass image. We illustrate the use of MIMS to image unlabeled mammalian cultured cells and tissue sections; to analyze fatty-acid transport in adipocyte lipid droplets using ^{13}C-oleic acid; to examine nitrogen fixation in bacteria using ^{15}N gaseous nitrogen; to measure levels of protein renewal in the cochlea and in post-ischemic kidney cells using ^{15}N-leucine; to study DNA and RNA co-distribution and uridine incorporation in the nucleolus using ^{15}N-uridine and ^{81}Br of bromodeoxyuridine or ^{14}C-thymidine; to reveal domains in cultured endothelial cells using the native isotopes ^{12}C, ^{16}O, ^{14}N and ^{31}P; and to track a few ^{15}N-labeled donor spleen cells in the lymph nodes of the host mouse. Conclusion: MIMS makes it possible for the first time to both image and quantify molecules labeled with stable or radioactive isotopes within subcellular compartments

Impact of triclosan on behaviour and neural development of Cyprinodon variegatus

The study focussed on the effects of triclosan (TCS) exposure on mobility and hearing capacities of Cyprinodon variegatus larvae. TCS is an omnipresent antimicrobial and contaminant of aquatic ecosystems, which can act as endocrine disruptor, mainly by modifying thyroid functioning. Larval stages are particularly vulnerable to deleterious effects of endocrine disrupters because of potential impairment of fish development and behaviour. Exposure to TCS was conducted at fertilization of eggs at concentrations likely to be found in the environment: 20, 50 and 100 μg.l-1. The analysis of growth parameters of C. variegatus showed no effect of TCS on the fertility of eggs, survival and larval weight. Subsequently, THs concentrations were measured on 15 days post hatching larvae. THs are initially produced as T4 (thyroxine) cells and then converted in the bioactive form of T3 (triiodothyronine) cells. The observed increase of T4 and T3 cells in larvae exposed to 50 and 100 μg.l-1 suggests an increase in THs synthesis as a consequence of TCS exposure. Auditory thresholds of larvae were determined using ABR (Auditory Brainstem Response) technique, and finally larval mobility was measured. For both parameters no significant differences were observed among the three different treatments. Audiograms showed that the auditory system is not yet completely established at 30 days post hatching. However, these results allowed us to consider C. variegatus as an “hearing generalist” because this species have a hearing sensitivity lower than 2000 Hz. Regarding locomotion, our result summarized short time experiences targeting only swimming speed, distance and degree of mobility. It would be interesting to expand the behavioural aspects on other parameters of locomotion and integrate Cyprinodon reaction to different stress (light or touch). In conclusion, our results require an extensive long-term study on the full life cycle of C. variegatus, in order to evaluate the impact of triclosan on neural function and behaviour through several generations

The thyroid gland and thyroid hormones in sheepshead minnow (Cyprinodon variegatus) during early development and metamorphosis

The sheepshead minnow is widely used in ecotoxicological studies that recently, begin to focus on potential disruption of the thyroid axis by xenobiotics and endocrine disrupting compounds. However, reference levels of thyroxin (T4) and 3,5,3’-triiodothyronine (T3) and their developmental patterns are unknown. This study set out to describe the ontogeny and morphology of the thyroid gland in sheepshead minnow, and correlate these with whole body concentrations of thyroid hormones during early development and metamorphosis. Couples of three females and two males were placed in breeding chambers designed for this experiment. More than 1000 eggs were collected and maintained in seawater. Embryos were selected under a dissection microscope and placed in incubation dishes (50 per dish) at 25°C. On day 6, embryos hatched and larvae were transferred to 1L beakers. For one week after hatching, larvae were fed on artemia, and from 8 to 30 days post-hatch they were fed on flaked fish food. Embryos were sampled on day 0, 3, 6 post-fertilization and larvae and juveniles were sampled every three days from day 0 to 28 days post-hatch. The pooled samples were taken from several incubation dishes and divided in three replicate batches of 10-30 individuals. T4 and T3 were extracted from whole fish bodies and an enzyme-linked immunoassay was used to measure whole-body hormone levels. At each sampling point 5 individuals were placed in formalin fixative for histology. Length and body mass were measured. Hatching success, gross morphology, thyroid hormone levels and histology data were recorded. The onset of metamorphosis at 12 days post-hatching coincided with surges in whole body T4 and T3 concentrations. Thyroid follicles were first observed in pre-metamorphic larvae at hatching, and were detected exclusively in the subpharyngeal region, surrounding the ventral aorta. Follicle size and epithelial cell heights varied during the developmental phase, indicating fluctuations in thyroid hormone synthesis activity. The increase in the whole body T3/T4 ratio was indicative of an increase in outer ring deiodination. This study establishes a baseline for thyroid hormones in sheepshead minnows, which will be vital for the understanding of thyroid hormone functions and in future studies of thyroid toxicants in this species

Evaluation of the impact of Triclosan on the functioning of the thyroid system in Cyprinodon variegatus L., 1803.

Triclosan (5-chloro-2-[2, 4-dichlorophenoxy] phenol) is an antimicrobial widely used in various industrial products such as textiles, cosmetics and body care products. It is often detected in aquatic environments. The presence of the main biotransformation product, methyl TCS, indicates that this compound is not only degraded, but also persistent and accumulates in aquatic organisms. In this study, the effects of TCS on the thyroid system during embryonic and larval stages in Cyprinodon variegatus were evaluated. In particular, whole body thyroid hormone levels and the activity of deiodinases, enzymes involved in the activation of the thyroid prohormone T4, were measured. Couples of three females and two males were placed in breeding chambers designed for this experiment from which 3832 eggs were obtained by reproduction in the laboratory. Eggs were collected and maintained in seawater. Embryos were selected under a dissection microscope, randomly assigned to each of five treatment groups: Control, DMSO control (vehicle), 20 μg/L TCS, 50 μg/L TCS and 100 μg/L TCS and placed in incubation dishes (50 per dish) at 25°C. On day 6, embryos hatched and larvae were transferred to 1L dishes. The larvae were fed on artemia and on flaked fish food till day 15 post hatching when the fish were analyzed. The fertilization and survival rate, as well as the larval dry weight did not vary significantly between individuals exposed to 20, 50 and 100 μg/L TCS. T3 and T4 concentrations increased significantly in larvae exposed to 50 and 100 μg/L TCS. The study of the activity of enzymes involved in the deiodination of thyroid hormones (ORD) represents a new aspect in the study of endocrine disruption in C. variegatus. Unfortunately, we were not able to detect a net enzymatic T4 deiodination activity, most likely due to the very small amounts of protein and low specific enzyme activity in brain homogenates, carcass and liver

THE EFFECTS OF TRICLOSAN ON HEARING DEVELOPMENT OF CYPRINODON VARIEGATUS LARVAE

The aquatic environment represents the final sink for many chemicals, including bactericidal agents. Among them Triclosan (TCS) has been shown to affect the thyroid system of teleost. Larval stages are particularly vulnerable to deleterious effects of endocrine disrupters because of potential impairment of fish development and behaviour. Thyroid hormones are critical to the development of the brain and auditory system. Thus, TCS could affect the development of the brain and hearing. The aims of this study were: to investigate hearing development in sheepshead minnows (Cyprinodon variegatus) using the ABR technique (Auditory Brainstem Response) and to investigate the effects of triclosan on hearing development. Exposure to TCS was conducted from fertilization of eggs on at concentrations likely to be found in the environment: 20, 50 and 100 μgl-1. We characterized previously the ontogenic variation of thyroid hormones in embryos and larvae of sheepshead minnows. We observed an increase of thyroid hormones level around the 12th and the 15th day post hatching (dph), that may be associated with the transition from larval to juvenile stage during the development of this species. We concluded, that this period could be defined as a critical exposure window to pollutants. We determined hearing thresholds for sheepshead minnows of different ages. Due to the small size of larvae, first measurements were only possible at 30 dph. Audiograms showed that the hearing ability is not yet completely established at 30 dph compared to adults. So we pursued our experiments in older larvae of 40, 60, 70, and 80 dph when sexual maturity is attained. First results indicate that the hearing ability considerably improves between 50 and 60 dph. The effects of TCS in this development have yet to be determined but will be fully discussed. This study proposes an interesting new endpoint in thyroid disruption research

Do Triclosan affect hearing development of Cyprinodon variegatus larvae?

The aquatic environment represents the final sink for many chemicals, including bactericidal agents. Among them Triclosan (TCS) has been shown to affect the thyroid system of teleost. Larval stages are particularly vulnerable to deleterious effects of endocrine disrupters because of potential impairment of fish development and behaviour. Thyroid hormones are critical to the development of the brain and auditory system. Thus, TCS could affect the development of the brain and hearing. The aims of this study were: to investigate hearing development in sheepshead minnows (Cyprinodon variegatus) using the ABR technique (Auditory Brainstem Response) and to investigate the effects of triclosan on hearing development. Exposure to TCS was conducted from fertilization of eggs on at concentrations likely to be found in the environment: 20, 50 and 100 µgl-1. We characterized previously the ontogenic variation of thyroid hormones in embryos and larvae of sheepshead minnows. We observed an increase of thyroid hormones level around the 12th and the 15th day post hatching (dph), that may be associated with the transition from larval to juvenile stage during the development of this species. We concluded, that this period could be defined as a critical exposure window to pollutants. We determined hearing thresholds for sheepshead minnows of different ages. Our sheepshead minnows show ontogenic variations in the hearing ability during their development. At 30 days post hatching, their hearing ability is quite bad, with a narrow bandwidth of detected frequencies. But their hearing ability considerably enhance during their development to reach the adult hearing ability at around 80 days post hatching when this species reach sexual maturity. So we observe during the developmental phase of this fish species clear ontogenic improvements of the hearing ability and they showed an ontogenetic expansion in the frequency bandwidth they were able to detect. The effects of TCS in this development have yet to be determined but will be fully discussed. This study proposes an interesting new endpoint in thyroid disruption research

Cell migration guided by long-lived spatial memory

International audienceLiving cells actively migrate in their environment to perform key biological functions-from unicellular organisms looking for food to single cells such as fibroblasts, leukocytes or cancer cells that can shape, patrol or invade tissues. Cell migration results from complex intracellular processes that enable cell self-propulsion, and has been shown to also integrate various chemical or physical extracellular signals. While it is established that cells can modify their environment by depositing biochemical signals or mechanically remodelling the extracellular matrix, the impact of such self-induced environmental perturbations on cell trajectories at various scales remains unexplored. Here, we show that cells can retrieve their path: by confining motile cells on 1D and 2D micropatterned surfaces, we demonstrate that they leave long-lived physicochemical footprints along their way, which determine their future path. On this basis, we argue that cell trajectories belong to the general class of self-interacting random walks, and show that self-interactions can rule large scale exploration by inducing long-lived ageing, subdiffusion and anomalous first-passage statistics. Altogether, our joint experimental and theoretical approach points to a generic coupling between motile cells and their environment, which endows cells with a spatial memory of their path and can dramatically change their space exploration

High-resolution quantitative imaging of mammalian and bacterial cells using stable isotope mass spectrometry

BACKGROUND: Secondary-ion mass spectrometry (SIMS) is an important tool for investigating isotopic composition in the chemical and materials sciences, but its use in biology has been limited by technical considerations. Multi-isotope imaging mass spectrometry (MIMS), which combines a new generation of SIMS instrument with sophisticated ion optics, labeling with stable isotopes, and quantitative image-analysis software, was developed to study biological materials. RESULTS: The new instrument allows the production of mass images of high lateral resolution (down to 33 nm), as well as the counting or imaging of several isotopes simultaneously. As MIMS can distinguish between ions of very similar mass, such as (12)C(15)N(- )and (13)C(14)N(-), it enables the precise and reproducible measurement of isotope ratios, and thus of the levels of enrichment in specific isotopic labels, within volumes of less than a cubic micrometer. The sensitivity of MIMS is at least 1,000 times that of (14)C autoradiography. The depth resolution can be smaller than 1 nm because only a few atomic layers are needed to create an atomic mass image. We illustrate the use of MIMS to image unlabeled mammalian cultured cells and tissue sections; to analyze fatty-acid transport in adipocyte lipid droplets using (13)C-oleic acid; to examine nitrogen fixation in bacteria using (15)N gaseous nitrogen; to measure levels of protein renewal in the cochlea and in post-ischemic kidney cells using (15)N-leucine; to study DNA and RNA co-distribution and uridine incorporation in the nucleolus using (15)N-uridine and (81)Br of bromodeoxyuridine or (14)C-thymidine; to reveal domains in cultured endothelial cells using the native isotopes (12)C, (16)O, (14)N and (31)P; and to track a few (15)N-labeled donor spleen cells in the lymph nodes of the host mouse. CONCLUSION: MIMS makes it possible for the first time to both image and quantify molecules labeled with stable or radioactive isotopes within subcellular compartments