11 research outputs found
The in vivo effect of chelidonine on the stem cell system of planarians
The presence of adult pluripotent stem cells and the amazing regenerative capabilities make planarian
flatworms an extraordinary experimental model to assess in vivo the effects of substances of both natural and
synthetic origin on stem cell dynamics. This study focuses on the effects of chelidonine, an alkaloid obtained
from Chelidonium majus. The expression levels of molecular markers specific for stem or differentiated cells
were compared in chelidonine-treated and control planarians. The use of these markers demonstrates that
chelidonine produces in vivo a significant anti-proliferative effect on planarian stem cells in a dosedependent
fashion. In response to chelidonine treatment mitotic abnormalities were also observed and the
number of cells able to proceed to anaphase/telophase appeared significantly reduced with respect to the
controls. Our results support the possibility that chelidonine acts on cell cycle progression by inhibition of
tubulin polymerization. These studies provide a basis for preclinical evaluation in vivo of the effects of
chelidonine on physiologically proliferating stem cells
Stem cell protection mechanisms in planarians: the role of some heat shock genes
Planarians contain a large population of stem cells, named neoblasts, and use these for continuous turnover of all cell types. In addition, thanks to the amazing flexibility of these cells, planarians respond well to the effects of stressful situations, for example activating regeneration after trauma. How neoblasts respond to stress and support continuous proliferation, maintaining long-term stability, is still an open question. Heat shock proteins (HSPs) are a complex protein family with key roles in maintaining protein homeostasis, as well as in apoptosis and growth-related processes. We recently characterized some planarian homologs of hsp genes that are highly expressed in mammalian stem cells, and observed that some of them are critical for neoblast survival/maintenance. The results of these studies support the notion that some HSPs play crucial roles in the modulation of pathways regulating stem cell activity, regeneration and tissue repair. In this review we compare the evidence available for planarian hsp genes and focus on questions emerging from these results
Expression of hsp90 mediates cytoprotective effects in the gastrodermis of planarians
Heat shock proteins (HSPs) play a crucial role in
the protection of cells. In the present study, we have identified
an hsp90-related gene (Djhsp90) encoding a cytosolic form
of HSP90 that is primarily expressed in gastrodermis of the
planarian Dugesia japonica. Djhsp90 becomes significantly
induced after traumatic amputation or other stress stimuli,
such as exposure to X-ray or ultraviolet radiations, heat
shock, or prolonged starvation. When Djhsp90 is silenced by
ribonucleic acid interference (RNAi), planarians dramatically
decrease in size, becoming unable to eat, and die in a few
weeks. Our results indicate that this gene plays an essential
cytoprotective role in the gastrodermis of planarians and
suggest that this chaperone can be involved in autophagic
processes that are activated by this tissue
Expression of hsp90 mediates cytoprotective effects in the gastrodermis of planarians
Heat shock proteins (HSPs) play a crucial role in the protection of cells. In the present study, we have identified an hsp90-related gene (Djhsp90) encoding a cytosolic form of HSP90 that is primarily expressed in gastrodermis of the planarian Dugesia japonica. Djhsp90 becomes significantly induced after traumatic amputation or other stress stimuli, such as exposure to X-ray or ultraviolet radiations, heat shock, or prolonged starvation. When Djhsp90 is silenced by ribonucleic acid interference (RNAi), planarians dramatically decrease in size, becoming unable to eat, and die in a few weeks. Our results indicate that this gene plays an essential cytoprotective role in the gastrodermis of planarians and suggest that this chaperone can be involved in autophagic processes that are activated by this tissue
Characterization of hsp genes in planarian stem cells
Planarians are a model system known for regenerative potential, body plasticity and continuous turnover of all differentiated cell types. These characteristics are based on the presence of pluripotent stem cells, called neoblasts. Damage or reduction in the number of neoblasts deeply affects planarian regeneration and survival. Heat shock proteins (HSPs) are known to perform essential cytoprotective functions in all organisms. To investigate the potential role of hsp-related genes on the dynamics of planarian stem cells, representative hsp-related genes were identified and characterized in normal conditions and after different stress stimuli. Our work revealed that two different hsp genes (Djhsp60 and Djmot) are constitutively expressed in neoblasts, suggesting that their products play important roles in cytoprotection of these cells. RNAi-based functional studies provide evidence of an involvement of Djhsp60 and Djmot in the adaptive response of planarian stem cells to stress and indicate that expression of these genes is critical for planarian survival
A mortalin-like gene is crucial for planarian stem cell viability
In adult organisms, stem cells are crucial to homeostasis and regeneration of damaged tissues. In planarians,
adult stem cells (neoblasts) are endowed with an extraordinary replicative potential that guarantees
unlimited replacement of all differentiated cell types and extraordinary regenerative ability. The molecular
mechanisms by which neoblasts combine long-term stability and constant proliferative activity, overcoming
the impact of time, remain by far unknown. Here we investigate the role of Djmot, a planarian orthologue
that encodes a peculiar member of the HSP70 family, named Mortalin, on the dynamics of stem cells of
Dugesia japonica. Planarian stem cells and progenitors constitutively express Djmot. Transient Djmot
expression in differentiated tissues is only observed after X-ray irradiation. DjmotRNA interference causes
inability to regenerate and death of the animals, as a result of permanent growth arrest of stem cells. These
results provide the first evidence that an hsp-related gene is essential for neoblast viability and suggest the
possibility that high levels of Djmot serve to keep a p53-like protein signaling under control, thus allowing
neoblasts to escape cell death programs. Further studies are needed to unravel the molecular pathways
involved in these processes
Characterization of hsp genes on planarian stem cells
Planarians are a model system known for regenerative potential, body plasticity and continuous turnover of all differentiated cell types. These characteristics are based on the presence of pluripotent stem cells, called neoblasts. Damage or reduction in the number of neoblasts deeply affects planarian regeneration and survival. Heat shock proteins (HSPs) are known to perform essential cytoprotective functions in all organisms. To investigate the potential role of hsp-related genes on the dynamics of planarian stem cells, representative hsp-related genes were identified and characterized in normal conditions and after different stress stimuli. Our work revealed that two different hsp genes (Djhsp60 and Djmot) are constitutively expressed in neoblasts, suggesting that their products play important roles in cytoprotection of these cells. RNAi-based functional studies provide evidence of an involvement of Djhsp60 and Djmot in the adaptive response of planarian stem cells to stress and indicate that expression of these genes is critical for planarian survival
Aiming to understand the mechanisms underlying the different responses to berberine among different cell lines
Berberine, a bioactive natural isoquinoline alkaloid, is known
to generate a variety of pharmacological effects in different cell
types. Because of its ability to arrest the cell cycle and cause
apoptosis in several malignant cell lines, berberine has received
attention as a potential anticancer agent. To investigate the
mechanisms underlying the different responses to berberine, we
started to analyze its dose-dependent and time-dependent intracellular
localization in two human tumor cell lines: MIA PaCa-
2 (from pancreatic carcinoma), U343 (from glioblastoma).
Human dermal fibroblasts (HDF) were used as a non-tumor
control. Berberine presents natural green fluorescence, which
allows identification of the intracellular site of accumulation in
living cells. We found that the alkaloid may accumulate in different
cell compartments, with a dynamic dose-dependent and
time-dependent pattern of localization. The results revealed different
localization of berberine in cytoplasm and mitochondria
and/or nuclei in cancer cells with respect to non-tumor cells.
Moreover, berberine treatments reduced cell viability in a cell
line-specific manner. To further investigate the effects of berberine,
the expression profile of genes involved at different levels in
fundamental biological processes, was analyzed. As tumor suppressor
genes are often methylated in the process of carcinogenesis,
we evaluated DNMT1 and DNMT3B coding for maintenance
and de novo methyltransferases, respectively.
Additionally, MGMT, a geneencoding O6-methylguanine-DNA
methyltransferase, and recognized to play a crucial role in the
defense against chemotherapy alkylating agents, was analyzed.
As literature data demonstrate that berberine may induce apoptosis
in cancer cells but not in normal cells and this observation
is valuable for development of new anti-cancer therapies, we
also compared the variation in the gene expression level of cysteine-
aspartic acid protease 3 (CASP3) and the activity of this
enzyme in MIA PaCa-2, U343 and HDF cell lines. On the whole
the results indicate that berberine differentially affects the behaviour of MIA PaCa-2, U343, and non-tumor HDF cells
Effects of natural compounds present in Chelidonium majus on stem cells and embryo development
Despite their potential attractiveness as model for regeneration, planarians have not been used yet in large-scale chemical screenings to test the effects of compounds on different key aspects of regeneration, such as stem cell proliferation and differentiation. Our work focuses on the analysis of the effects produced by the main alkaloids (chelidonine, berberine, sanguinarine) present in Chelidonium majus, an herb with therapeutical properties. The data obtained so far indicate that chelidonine generates anti-proliferative effects on planarian stem cells, possibly due to inhibition of tubulin polymerization, berberine induces a perturbation of the regenerative pattern and sanguinarine produces abnormal head regeneration. In particular, we find that berberine causes abnormal regeneration of the visual system, without affecting cell proliferation/apoptosis, while sanguinarine induces apoptosis through a caspase dependent mechanism. These results clearly indicate the potentiality of planarians as a model to analyze drug effects. We are currently exploiting the well-established zebrafish model to assess and compare the effects of the same compounds during vertebrate embryo development