ROLE OF THE MESENCHYME IN THE INDUCTION OF THE RAT PROSTATE GLAND BY ANDROGENS IN ORGAN CULTURE

1979 ◽  
Vol 82 (1) ◽  
pp. 171-NP ◽  
Author(s):  
ILSE LASNITZKI ◽  
TAKEO MIZUNO
Keyword(s):  
Organ Culture ◽  
De Novo ◽  
Prostate Gland ◽  
Young Male ◽  
Free Medium ◽  
Foetal Rat ◽  
Rat Prostate ◽  
Do So

SUMMARY Rat prostate glands are induced de novo by androgens in 16·5-day-old male and female urogenital sinuses in vitro as epithelial buds projecting into the surrounding mesenchyme. The role of the mesenchyme in this process has been investigated in various epithelial-mesenchymal recombinations in organ culture. Isolated epithelium did not form buds but required the presence of the mesenchyme to do so. This requirement seemed to be specific; in the presence of testosterone or dihydrotestosterone only urogenital mesenchyme increased cell division in the urogenital epithelium and stimulated prostatic bud formation. In contrast, heterotypic mesenchyme did not affect epithelial mitosis and failed to induce buds while heterotypic epithelia did not respond to urogenital mesenchyme. In recombinants of urogenital mesenchyme pretreated with androgen and untreated urogenital epithelium, grown in androgen-free medium, the majority of explants developed prostatic buds while only a few buds were formed from epithelium pretreated with androgen when it was recombined with untreated mesenchyme. The role of the mesenchyme in the loss of androgen responsiveness of the older female sinuses was examined in heterochronic recombinants. It was found that the old female mesenchyme failed to induce buds in young epithelium while young male or female mesenchymes induced them in the old female epithelium. The results suggest that the urogenital mesenchyme is essential for the initiation of the foetal rat prostate gland and that it may be a target for androgens and complement or mediate their effect on the epithelium.

INDUCTION OF THE RAT PROSTATE GLAND BY ANDROGENSIN ORGAN CULTURE

1977 ◽  
Vol 74 (1) ◽  
pp. 47-55 ◽  
Author(s):  
ILSE LASNITZKI ◽  
TAKEO MIZUNO
Keyword(s):  
Organ Culture ◽  
De Novo ◽  
Close Contact ◽  
Prostate Gland ◽  
Wistar Rat ◽  
Continuous Treatment ◽  
Control Medium ◽  
Androgenic Activity ◽  
Prostatic Epithelium ◽  
Rat Prostate

The induction of the prostate gland by testosterone or dihydrotestosterone was studied in urogenital sinuses from Wistar rat embryos explanted in organ culture. In normal development, prostatic rudiments appeared in the urogenital sinuses of 18–19 day male foetuses as epithelial buds projecting into the mesenchyme. Male urogenital sinuses from 17·5 and 18·5 day foetuses formed buds in control medium, and addition of testosterone or dihydrotestosterone increased their number and size. In contrast, sinuses from 15·5 and 16·5 day male foetuses did not form buds in the absence of androgens, while exposure to the hormones induced them de novo. A concentration of 0·0015 μg testosterone/ml was sufficient to elicit prostatic buds. The same effect was seen after continuous treatment with testosterone or brief exposure to the hormone followed by cultivation in control medium. The development of the buds followed the same time sequence as in the organ in situ. Testosterone and dihydrotestosterone elicited prostatic buds in female sinuses from 15·5 to 18·5 day foetuses, but their response to androgens decreased with advancing foetal age. Male and female sinuses from 14·5 day foetuses did not respond to testosterone but formed buds after exposure to dihydrotestosterone. This difference was considered to be due to a low content or absence of 5α-reductase at this stage. Testes from 15·5 to 18·5 day foetuses grown in close contact with urogenital sinuses or explants of mature prostate glands induced prostatic buds or augmented the height of the prostatic epithelium. It was concluded that androgens are necessary for the initiation of the rat prostate gland and that the effect of testosterone is mediated via 5α-dihydrotestosterone. The onset of 5α-reductase synthesis is likely to occur between days 14 and 15 of foetal life. The early androgenic activity of the testes suggests that bud formation in the older male sinuses in the absence of androgens is due to exposure to endogenous testosterone before explantation.

THE BINDING OF [1,2-3H]TESTOSTERONE WITHIN NUCLEI OF THE RAT PROSTATE

1969 ◽  
Vol 44 (3) ◽  
pp. 323-333 ◽  
Author(s):  
W. I. P. MAINWARING
Keyword(s):  
Molecular Weight ◽  
Equilibrium Dialysis ◽  
Prostate Gland ◽  
Ventral Prostate ◽  
Rat Tissues ◽  
Receptor Complex ◽  
Rat Prostate ◽  
Binding Component

SUMMARY The specificity of the binding of [1,2-3H]testosterone to nuclei of various rat tissues in vivo has been studied. A significant amount of radioactivity was retained in the nuclei of androgen-dependent tissues only, particularly the ventral prostate gland. The bound radioactivity was only partially recovered as [1,2-3H]testosterone; the remainder was identified as [3H]5α-dihydrotestosterone. Efforts were made to characterize the binding component, or 'receptor', in prostatic nuclei. On digestion of nuclei labelled in vivo with [1,2-3H]testosterone, with enzymes of narrow substrate specificity, only trypsin released tritium, suggesting that the receptor is a protein. On the basis of subfractionation studies of labelled nuclei, the receptor is an acidic protein. The androgen—receptor complex could be effectively extracted from the prostatic nuclei in 1 m-NaCl and from the results of fractionations on a calibrated agarose column, the complex has a molecular weight 100,000–120,000. The specificity of the binding of steroids to such 1 m-NaCl extracts in vitro was investigated by the equilibrium dialysis procedure. Under these conditions, the specificity of the binding of [1,2-3H]testosterone demonstrated in vivo could not be simulated. The receptor is probably part of the chromatin complex but its precise intranuclear localization cannot be determined by biochemical procedures alone.

scholarly journals Cytoplasmic Expression of the ALS/FTD-Related Protein TDP-43 Decreases Global Translation Both in vitro and in vivo

2020 ◽  
Vol 14 ◽  
Author(s):  
Santiago E. Charif ◽  
Luciana Luchelli ◽  
Antonella Vila ◽  
Matías Blaustein ◽  
Lionel M. Igaz
Keyword(s):  
De Novo ◽  
Brain Slices ◽  
Mrna Translation ◽  
Hek293 Cells ◽  
Cytoplasmic Inclusions ◽  
Cytoplasmic Expression ◽  
Global Translation

TDP-43 is a major component of cytoplasmic inclusions observed in neurodegenerative diseases like frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). To further understand the role of TDP-43 in mRNA/protein metabolism and proteostasis, we used a combined approach with cellular and animal models overexpressing a cytoplasmic form of human TDP-43 (TDP-43-ΔNLS), recapitulating ALS/FTD features. We applied in HEK293 cells a method for labeling de novo translation, surface sensing of translation (SUnSET), based on puromycin (PURO) incorporation. While control cells displayed robust puromycilation, TDP-43-ΔNLS transfected cells exhibited reduced ongoing protein synthesis. Next, by using a transgenic mouse overexpressing cytoplasmic TDP-43 in the forebrain (TDP-43-ΔNLS mice) we assessed whether cytoplasmic TDP-43 regulates global translation in vivo. Polysome profiling of brain cortices from transgenic mice showed a shift toward non-polysomal fractions as compared to wild-type littermates, indicating a decrease in global translation. Lastly, cellular level translational assessment by SUNSET was performed in TDP-43-ΔNLS mice brain slices. Control mice slices incubated with PURO exhibited robust cytoplasmic PURO signal in layer 5 neurons from motor cortex, and normal nuclear TDP-43 staining. Neurons in TDP-43-ΔNLS mice slices incubated with PURO exhibited high cytoplasmic expression of TDP-43 and reduced puromycilation respect to control mice. These in vitro and in vivo results indicate that cytoplasmic TDP-43 decreases global translation and potentially cause functional/cytotoxic effects as observed in ALS/FTD. Our study provide in vivo evidence (by two independent and complementary methods) for a role of mislocalized TDP-43 in the regulation of global mRNA translation, with implications for TDP-43 proteinopathies.

scholarly journals LINC01006 facilitates cell proliferation, migration and invasion in prostate cancer through targeting miR-34a-5p to up-regulate DAAM1

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Enhui Ma ◽  
Qianqian Wang ◽  
Jinhua Li ◽  
Xinqi Zhang ◽  
Zhenjia Guo ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Prostate Gland ◽  
Inhibitory Effect ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Protein Coding ◽  
Novel Target

Abstract Background Prostate cancer (PCa) is a kind of malignancy occurring in the prostate gland. Substantial researches have proved the major role of long noncoding RNAs (lncRNAs) in PCa. However, the role of long intergenic non-protein coding RNA 1006 (LINC01006) in PCa has not been investigated yet. Methods RT-qPCR was used to examine the expression levels of LINC01006 and its downstream targets. The function of LINC01006 in PCa was tested by in vitro and in vivo assays. With application of RNA pull down, RNA immunoprecipitation (RIP) and luciferase reporter assays, the interaction among LINC01006, miR-34a-5p and disheveled associated activator of morphogenesis 1 (DAAM1) were verified. Results LINC01006 expression presented high in PCa cell lines. LINC01006 silencing suppressed cell proliferative, migratory, invasive capacities while accelerated apoptotic rate. Besides, LINC01006 knockdown also suppressed tumor growth and metastasis in vivo. Furthermore, miR-34a-5p, a tumor suppressor in PCa, was sponged by LINC01006. Moreover, DAAM1 was targeted by miR-34a-5p and promoted PCa progression. More intriguingly, rescue assays suggested that the inhibitory effect of LINC01006 knockdown on PCa development was offset by DAAM1 overexpression. Conclusions LINC01006 promoted PCa progression by sponging miR-34a-5p to up-regulate DAAM1, providing a novel target for PCa therapy.

Possible role of gap junctions in activation of myometrium during parturition

1978 ◽  
Vol 235 (5) ◽  
pp. C168-C179 ◽  
Author(s):  
R. E. Garfield ◽  
S. M. Sims ◽  
M. S. Kannan ◽  
E. E. Daniel
Keyword(s):  
Smooth Muscle ◽  
Gap Junctions ◽  
Freeze Fracture ◽  
Normal Pregnancy ◽  
Pregnant Rats ◽  
Junction Formation ◽  
Synchronous Contraction ◽  
Do So

Gap junctions between smooth muscle cells of the myometrium of pregnant rats were found only immediately prior to, during and immediately after parturition by quantitative thin-section and freeze-fracture microscopy. Ovariectomy of 16- to 17-days-pregnant rats resulted in premature termination of pregnancy and the appearance of gap junctions. Methods that prolonged normal pregnancy in rats or maintained pregnancy in ovariectomized animals (progesterone treatment) prevented the appearance of gap junctions. Gap junctions formed in tissues incubated for 24--96 h in vitro without any hormonal influence. We propose that gap junctions are essential for normal labor and delivery for synchronous contraction of the muscle of the uterus. We present a model for control of parturition that may apply to other animals including humans. The model proposes: 1) the possible roles progesterone, prostaglandins, or estrogens may play in initiating gap-junction formation; 2) that the formation of gap junctions is a necessary step in activation of the myometrium leading to labor; and 3) that agents used to stimulate or inhibit labor may do so by affecting gap junctions.

scholarly journals NAD+-Dependent Deacetylase Hst1p Controls Biosynthesis and Cellular NAD+ Levels in Saccharomyces cerevisiae

2003 ◽  
Vol 23 (19) ◽  
pp. 7044-7054 ◽  
Author(s):  
Antonio Bedalov ◽  
Maki Hirao ◽  
Jeffrey Posakony ◽  
Melisa Nelson ◽  
Julian A. Simon
Keyword(s):  
De Novo ◽  
Critical Role ◽  
Salvage Pathway ◽  
De Novo Synthesis ◽  
Array Analysis ◽  
Binding Partner ◽  
Dependent Protein ◽  
New Protein

ABSTRACT Nicotine adenine dinucleotide (NAD+) performs key roles in electron transport reactions, as a substrate for poly(ADP-ribose) polymerase and NAD+-dependent protein deacetylases. In the latter two processes, NAD+ is consumed and converted to ADP-ribose and nicotinamide. NAD+ levels can be maintained by regeneration of NAD+ from nicotinamide via a salvage pathway or by de novo synthesis of NAD+ from tryptophan. Both pathways are conserved from yeast to humans. We describe a critical role of the NAD+-dependent deacetylase Hst1p as a sensor of NAD+ levels and regulator of NAD+ biosynthesis. Using transcript arrays, we show that low NAD+ states specifically induce the de novo NAD+ biosynthesis genes while the genes in the salvage pathway remain unaffected. The NAD+-dependent deacetylase activity of Hst1p represses de novo NAD+ biosynthesis genes in the absence of new protein synthesis, suggesting a direct effect. The known Hst1p binding partner, Sum1p, is present at promoters of highly inducible NAD+ biosynthesis genes. The removal of HST1-mediated repression of the NAD+ de novo biosynthesis pathway leads to increased cellular NAD+ levels. Transcript array analysis shows that reduction in cellular NAD+ levels preferentially affects Hst1p-regulated genes in comparison to genes regulated with other NAD+-dependent deacetylases (Sir2p, Hst2p, Hst3p, and Hst4p). In vitro experiments demonstrate that Hst1p has relatively low affinity toward NAD+ in comparison to other NAD+-dependent enzymes. These findings suggest that Hst1p serves as a cellular NAD+ sensor that monitors and regulates cellular NAD+ levels.

scholarly journals Identification of a talin binding site in the cytoskeletal protein vinculin.

1989 ◽  
Vol 109 (6) ◽  
pp. 2917-2927 ◽  
Author(s):  
P Jones ◽  
P Jackson ◽  
G J Price ◽  
B Patel ◽  
V Ohanion ◽  
...  
Keyword(s):  
Amino Acids ◽  
In Vitro Transcription ◽  
Binding Activity ◽  
Cytoskeletal Protein ◽  
Translation System ◽  
Cos Cells ◽  
Cell Matrix ◽  
Do So

Binding of the cytoskeletal protein vinculin to talin is one of a number of interactions involved in linking F-actin to cell-matrix junctions. To identify the talin binding domain in vinculin, we expressed the NH2-terminal region of the molecule encoded by two closely similar, but distinct vinculin cDNAs, using an in vitro transcription translation system. The 5' Eco RI-Bam HI fragment of a partial 2.89-kb vinculin cDNA encodes a 45-kD polypeptide containing the first 398 amino acids of the molecule. The equivalent restriction enzyme fragment of a second vinculin cDNA (cVin5) lacks nucleotides 746-867, and encodes a 41-kD polypeptide missing amino acids 167-207. The radiolabeled 45-kD vinculin polypeptide bound to microtiter wells coated with talin, but not BSA, and binding was inhibited by unlabeled vinculin. In contrast, the 41-kD vinculin polypeptide was devoid of talin binding activity. The role of residues 167-207 in talin binding was further analyzed by making a series of deletions spanning this region, each deletion of seven amino acids contiguous with the next. Loss of residues 167-173, 174-180, 181-187, 188-194, or 195-201 resulted in a marked reduction in talin binding activity, although loss of residues 202-208 had much less effect. When the 45-kD vinculin polypeptide was expressed in Cos cells, it localized to cell matrix junctions, whereas the 41-kD polypeptide, lacking residues 167-207, was unable to do so. Interestingly, some deletion mutants with reduced ability to bind talin in vitro, were still able to localize to cell matrix junctions.

The Role of Nucleophosmin In Hematopoietic Stem Cells and the Pathogenesis of Myelodysplastic Syndrome

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 95-95 ◽  
Author(s):  
Keisuke Ito ◽  
Paolo Sportoletti ◽  
John G Clohessy ◽  
Grisendi Silvia ◽  
Pier Paolo Pandolfi
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Myelodysplastic Syndrome ◽  
Cell Biology ◽  
De Novo ◽  
Stem Cell Biology ◽  
Hematopoietic Stem

Abstract Abstract 95 Myelodysplastic syndrome (MDS) is an incurable stem cell disorder characterized by ineffective hematopoiesis and an increased risk of leukemia transformation. Nucleophosmin (NPM) is directly implicated in primitive hematopoiesis, the pathogenesis of hematopoietic malignancies and more recently of MDS. However, little is known regarding the molecular role and function of NPM in MDS pathogenesis and in stem cell biology. Here we present data demonstrating that NPM plays a critical role in the maintenance of hematopoietic stem cells (HSCs) and the transformation of MDS into leukemia. NPM is located on chromosome 5q and is frequently lost in therapy-related and de novo MDS. We have previously shown that Npm1 acts as a haploinsufficient tumor suppressor in the hematopoietic compartment and Npm1+/− mice develop a hematologic syndrome with features of human MDS, including increased susceptibility to leukemogenesis. As HSCs have been demonstrated to be the target of the primary neoplastic event in MDS, a functional analysis of the HSC compartment is essential to understand the molecular mechanisms in MDS pathogenesis. However, the role of NPM in adult hematopoiesis remains largely unknown as Npm1-deficiency leads to embryonic lethality. To investigate NPM function in adult hematopoiesis, we have generated conditional knockout mice of Npm1, using the Cre-loxP system. Analysis of Npm1 conditional mutants crossed with Mx1-Cre transgenic mice reveals that Npm1 plays a crucial role in adult hematopoiesis and ablation of Npm1 in adult HSCs leads to aberrant cycling and followed by apoptosis. Analysis of cell cycle status revealed that HSCs are impaired in their ability to maintain quiescence after Npm1-deletion and are rapidly depleted in vivo as well as in vitro. Competitive reconstitution assay revealed that Npm1 acts cell-autonomously to maintain HSCs. Conditional inactivation of Npm1 leads to an MDS phenotype including a profoundly impaired ability to differentiate into cells of the erythroid lineage, megakaryocyte dyspoiesis and centrosome amplification. Furthermore, Npm1 loss evokes a p53-dependent response and Npm1-deleted HSCs undergo apoptosis in vivo and in vitro. Strikingly, transfer of the Npm1 mutation into a p53-null background rescued the apoptosis of Npm1-ablated HSCs and resulted in accelerated transformation to an aggressive and lethal form of acute myeloid leukemia. Our findings highlight the crucial role of NPM in stem cell biology and identify a new mechanism by which MDS can progress to leukemia. This has important therapeutic implications for de novo MDS as well as therapy-related MDS, which is known to rapidly evolve to leukemia with frequent loss or mutation of TRP53. Disclosures: No relevant conflicts of interest to declare.

Role of polyamines on de novo shoot morphogenesis from cotyledons of Brassica campestris ssp. pekinensis (Lour) Olsson in vitro

1994 ◽  
Vol 13 (6) ◽  
Author(s):  
Gek-Lan Chi ◽  
Wen-Shu Lin ◽  
JustinE.E. Lee ◽  
Eng-Chong Pua
Keyword(s):  
Brassica Campestris ◽  
De Novo ◽  
Shoot Morphogenesis
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