scholarly journals Effects of reproductive stage, GH, and 11-ketotestosterone on expression of growth differentiation factor-9 in the ovary of the eel, Anguilla australis

Reproduction ◽  
2010 ◽  
Vol 139 (1) ◽  
pp. 71-83 ◽  
Author(s):  
P M Lokman ◽  
Y Kazeto ◽  
Y Ozaki ◽  
S Ijiri ◽  
R Tosaka ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Mrna Levels ◽  
Gh Treatment ◽  
Oocyte Growth ◽  
Anguilla Australis ◽  
Differentiation Factor ◽  
Growth Differentiation Factor 9 ◽  
Pituitary Homogenate

In order to study the regulation of the growth differentiation factor-9 (gdf9) gene in a primitive teleost with semelparous life history, we cloned a cDNA encoding shortfinned eel Gdf9, expressed a partial peptide inEscherichia coli, and raised an antiserum to evaluate changes in Gdf9 expression during its pituitary homogenate-induced reproductive cycle. The effects ofin vivoandin vitroexposure to the androgen 11-ketotestosterone (11-KT), known to affect previtellogenic (PV) oocyte growth, were also determined. Furthermore, we investigated whether Gdf9 expression was metabolically gated by treating PV fish with recombinant GHin vivo. Immunoreactive proteins ofca. 52 and 55 kDa were identified by western blot analysis. Gdf9 message and protein were most abundant in PV oocytes, and peaked slightly earlier for mRNA than for protein. Captivity resulted in reducedgdf9mRNA levels, which were restored following pituitary homogenate treatment. As oocytes progressed through induced oogenesis, Gdf9 expression decreased. Neither 11-KT nor GH treatment affectedgdf9mRNA levels in PV fish, although GH could partially restore handling- or captivity-induced decreases ingdf9mRNA levels. Semelparous eels thus show an expression pattern of Gdf9 during oogenesis that is similar to that seen in other vertebrates, that appears responsive to handling or captivity stress, and whose control remains to be elucidated.

Effects of growth differentiation factor-9 and FSH on in vitro development, viability and mRNA expression in bovine preantral follicles

2013 ◽  
Vol 25 (8) ◽  
pp. 1194 ◽  
Author(s):  
G. L. Vasconcelos ◽  
M. V. A. Saraiva ◽  
J. J. N. Costa ◽  
M. J. Passos ◽  
A. W. B. Silva ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Nuclear Antigen ◽  
Mrna Levels ◽  
Related Factors ◽  
Differentiation Factor ◽  
Growth Differentiation Factor 9 ◽  
Before And After ◽  
Vitro Development

The present study investigated the role of growth differentiation factor (GDF)-9 and FSH, alone or in combination, on the growth, viability and mRNA expression of FSH receptor, proliferating cell nuclear antigen (PCNA) and proteoglycan-related factors (i.e. hyaluronan synthase (HAS) 1, HAS2, versican, perlecan) in bovine secondary follicles before and after in vitro culture. After 12 days culture, sequential FSH (100 ng mL–1 from Days 0 to 6 and 500 ng mL–1 from Days 7 to 12) increased follicular diameter and resulted in increased antrum formation (P < 0.05). Alone, 200 ng mL–1 GDF-9 significantly reduced HAS1 mRNA levels, but increased versican and perlecan mRNA levels in whole follicles, which included the oocyte, theca and granulosa cells. Together, FSH and GDF-9 increased HAS2 and versican (VCAN) mRNA levels, but decreased PCNA mRNA expression, compared with levels in follicles cultured in α-minimum essential medium supplemented with 3.0 mg mL–1 bovine serum albumin, 10 µg mL–1 insulin, 5.5 µg mL–1 transferrin, 5 ng mL–1 selenium, 2 mM glutamine, 2 mM hypoxanthine and 50 μg mL–1 ascorbic acid (α-MEM+). Comparisons of uncultured (0.2 mm) and α-MEM+ cultured follicles revealed that HAS1 mRNA expression was higher, whereas VCAN expression was lower, in cultured follicles (P < 0.05). Expression of HAS1, VCAN and perlecan (HSPG2) was higher in cultured than in vivo-grown (0.3 mm) follicles. In conclusion, FSH and/or GDF-9 promote follicular growth and antrum formation. Moreover, GDF-9 stimulates expression of versican and perlecan and interacts positively with FSH to increase HAS2 expression.

270. Androgens augment the mitogenic effects of oocyte-secreted factors and growth differentiation factor 9 on porcine granulosa cells

2005 ◽  
Vol 17 (9) ◽  
pp. 111
Author(s):  
T. E. Hickey ◽  
D. L. Marrocco ◽  
F. Amato ◽  
L. J. Ritter ◽  
R. J. Norman ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
Tritiated Thymidine ◽  
Paracrine Signalling ◽  
Secreted Factors ◽  
Differentiation Factor ◽  
Porcine Granulosa Cells ◽  
Growth Differentiation Factor 9 ◽  
Growth Promoting

Androgens, acting directly through the androgen receptor (AR), are thought to promote granulosa cell (GC) growth in vivo, but generally inhibit growth and promote GC differentiation in vitro. We hypothesised that the growth-promoting action of androgens on GC requires paracrine signalling from the oocyte. To test this hypothesis, we cultured mural GC from small antral (1–3mm) pre-pubertal pig follicles in the presence or absence of denuded oocytes (DO) from the same follicles to examine whether mitogenic responses, determined by uptake of tritiated thymidine, to combinations of FSH, insulin like growth factor 1 (IGF1) and dihydrotestosterone (DHT; 500 nM), were influenced by oocyte-secreted factors (OSFs). To further explore the identity of such factors, we performed the same experiments, substituting recombinant mouse growth differentiation factor 9 (GDF9), a known OSF, for the DO. Alone, DHT induced a small (<2-fold), but consistent increase in IGF1-stimulated DNA synthesis. OSFs stimulated DNA synthesis in all experimental combinations, most significantly in the presence of IGF1 (P < 0.0001), and DHT enhanced (P<0.05) the stimulatory effect of OSFs in all instances. Like OSFs, GDF9 substantially increased IGF1-stimulated DNA synthesis (P < 0.0001), and again, DHT enhanced (P > 0.01) this effect. In further experiments, two AR agonists, testosterone (10-1000nM) and DHT (5–500 nM), dose-dependently augmented the mitogenic effect of OSFs or GDF9 in the presence of IGF1. Only the highest doses of androgen had an independent stimulatory effect; lower doses required the presence of an OSF(s). Antiandrogen (hydroxyflutamide) treatment, used to block AR activity, antagonized the androgen X GDF9 interaction. In conclusion, androgens, via activation of the AR, stimulate porcine GC proliferation in vitro by potentiating the growth-promoting effects of oocytes or GDF9. These signalling pathway interactions are likely to be important regulators of folliculogenesis in vivo and may cause the excess follicle growth that is observed in androgen-treated female animals.

scholarly journals Growth Differentiation Factor 9 Enhances Activin A-Induced Inhibin B Production in Human Granulosa Cells

Endocrinology ◽  
2009 ◽  
Vol 150 (8) ◽  
pp. 3540-3546 ◽  
Author(s):  
Feng-Tao Shi ◽  
Anthony P. Cheung ◽  
Peter C. K. Leung
Keyword(s):  
Signaling Pathway ◽  
Mrna Level ◽  
Activin A ◽  
Inhibin B ◽  
Mrna Levels ◽  
Vitro Fertilization ◽  
Differentiation Factor ◽  
Protein Receptor ◽  
Growth Differentiation Factor 9

Activin A or growth differentiation factor 9 (GDF9) alone can increase βB-mRNA level in human granulosa-lutein cells from women undergoing in vitro fertilization, but their potential interactions and related cell signaling pathways involved are unknown. We therefore compared inhibin subunit and inhibin levels and activation of activin receptors (ACVRs) and Smad signaling pathway in these human granulosa-lutein cells with and without GDF9 and/or activin A treatment. Inhibin subunit (α, βA, βB), ACVR, and Smad2/3/4/7 mRNA levels, inhibin A and B production, and Smad phosphorylation were assessed by real-time RT-PCR, ELISA, and immunoblotting, respectively. Data were analyzed by ANOVA followed by Tukey’s test. Activin A (1–50 ng/ml) or GDF9 (1–200 ng/ml) alone had only little stimulatory effects on α- and βA-mRNA levels. In contrast, GDF9 could stimulate βB-subunit levels but to a lesser degree than the dose- and time-dependent effects of activin A. Compared with untreated cells, GDF9 pretreatment for 24 h significantly enhanced activin A-induced βB-mRNA levels, inhibin B secretion, and Smad2/3 phosphorylation (effects attenuated by bone morphogenetic protein receptor 2 extracellular domain, a GDF9 antagonist); and induced ACVR2B/1B and Smad2/3 but reduced Smad7 (an inhibitory Smad) mRNA levels. We report here for the first time that GDF9 enhances cell response to activin A by modulating key components of the activin signaling pathway in regulating inhibin subunits and hence inhibin B production in human granulosa-lutein cells.

scholarly journals Expression of growth differentiation factor 9, bone morphogenetic protein 15, and anti-Müllerian hormone in cultured mouse primary follicles

Reproduction ◽  
2008 ◽  
Vol 136 (2) ◽  
pp. 195-203 ◽  
Author(s):  
J C Sadeu ◽  
T Adriaenssens ◽  
J Smitz
Keyword(s):  
Bone Morphogenetic Protein ◽  
Neonatal Mouse ◽  
Mrna Levels ◽  
Differentiation Factor ◽  
Morphogenetic Protein ◽  
Secondary Stage ◽  
Growth Differentiation Factor 9 ◽  
Protein Expressions ◽  
Bone Morphogenetic Protein 15

Growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15), and anti-Müllerian hormone (AMH) play an important role in the primary to secondary follicle transition and follicle activation in vivo. In organ culture of neonatal mouse ovaries, it was observed that significantly fewer primary follicles develop to the secondary stage. The objectives of this study were: (1) to compare ovarian follicular populations between organ-cultured neonatal mouse ovaries and freshly isolated age-matched control ovaries; (2) to quantify RNA levels of Gdf9, Bmp15, and Amh in cultured primary follicles; and (3) to immunolocalize GDF9 and AMH in cultured ovaries. Ovaries from 3-day-old (PND 3) mice were cultured for 7 or 10 days in the absence or presence of FSH. Follicular populations were counted in freshly isolated 13-day-old (PND 13) ovaries and organ-cultured ovaries. Transcripts were quantified in isolated primary follicles using real-time RT-PCR, and protein expressions were localized using immunohistochemistry. The number of secondary follicles in organ-cultured ovaries was significantly lower than in vivo controls. Gdf9 and Bmp15 mRNA expression levels were similar as in controls. Amh mRNA levels were significantly (P<0.05) lower after day 10 of culture in the absence of FSH. GDF9 and AMH proteins were respectively detected in the oocytes and the granulosa cells (GC) beginning at the primary and primordial stages onward. GDF9 and BMP15 production in cultured primary follicles are not different from in vivo controls; hence abnormal early follicular growth was not related to a deficient transcription of these factors.

Безопасность соединений из группы терпенов ментанового ряда in vitro и in vivo

2020 ◽  
Vol 83 (4) ◽  
pp. 24-30
Author(s):  
Ирина Владимировна Акулина ◽  
Светлана Ивановна Павлова ◽  
Ирина Семеновна Степаненко ◽  
Назира Сунагатовна Карамова ◽  
Александр Владиславович Сергеев ◽  
...  
Keyword(s):  
Escherichia Coli

Проведено токсикологическое исследование соединений с антибактериальными свойствами из группы терпенов ментанового ряда в условиях in vitro и in vivo: лимонена (B34), его производного (+)-1,2-оксида лимонена (B60) и серосодержащего монотерпенового соединения 2-(1’-гидрокси-4’-изопренил-1’-метилциклогексил-2’-тио)метилэтаноата (B65). В условиях in vitro (культура опухолевых клеток HeLa) изучаемые монотерпены в диапазоне концентраций 2 – 200 мкг/мл обладали цитотоксичностью. Ингибирующая концентрация (ИК50) для B34 составила 231 (167 – 295) мкг/мл, для B60 – 181 (105 – 257) мкг/мл, ИК50 B65 – 229 (150 – 308) мкг/мл. Исследование генотоксичности показало, что B34 и B65 в диапазоне концентраций 50 – 1000 мкг/мл не индуцируют SOS мутагенез в клетках Escherichia coli PQ37, тогда как B60 в концентрациях 500 и 1000 мкг/мл проявляет генотоксичность. In vivo в остром эксперименте на беспородных мышах установлена низкая токсичность B34 и его производных при различных путях введения. Наименьший показатель острой токсичности имеет B65, в связи с чем дополнительно на крысах проведено изучение его хронической токсичности. Ежедневное внутрижелудочное введение B65 в разовых дозах, составляющих 1/10 и 1/20 ЛД50 (1000 мг/кг и 500 мг/кг), в течение 1 мес не вызывало гибели животных, значимых нарушений общего состояния, изменения динамики массы тела, морфопатологических изменений. Внутрижелудочное введение B65 крысам в высокой токсической дозе 2000 мг/кг (1/5 ЛД50) в течение месяца вызывает патоморфологические изменения структуры печени.

Nanocurcumin: A Double-Edged Sword for Microcancers

2020 ◽  
Vol 26 (45) ◽  
pp. 5783-5792
Author(s):  
Kholood Abid Janjua ◽  
Adeeb Shehzad ◽  
Raheem Shahzad ◽  
Salman Ul Islam ◽  
Mazhar Ul Islam
Keyword(s):  
Intracellular Signaling ◽  
Dosage Forms ◽  
The Body ◽  
In Vivo Studies ◽  
Mrna Levels ◽  
Anticancer Activities ◽  
Road Map ◽  
Anticancer Effects

There is compelling evidence that drug molecules isolated from natural sources are hindered by low systemic bioavailability, poor absorption, and rapid elimination from the human body. Novel approaches are urgently needed that could enhance the retention time as well as the efficacy of natural products in the body. Among the various adopted approaches to meet this ever-increasing demand, nanoformulations show the most fascinating way of improving the bioavailability of dietary phytochemicals through modifying their pharmacokinetics and pharmacodynamics. Curcumin, a yellowish pigment isolated from dried ground rhizomes of turmeric, exhibits tremendous pharmacological effects, including anticancer activities. Several in vitro and in vivo studies have shown that curcumin mediates anticancer effects through the modulation (upregulation and/or downregulations) of several intracellular signaling pathways both at protein and mRNA levels. Scientists have introduced multiple modern techniques and novel dosage forms for enhancing the delivery, bioavailability, and efficacy of curcumin in the treatment of various malignancies. These novel dosage forms include nanoparticles, liposomes, micelles, phospholipids, and curcumin-encapsulated polymer nanoparticles. Nanocurcumin has shown improved anticancer effects compared to conventional curcumin formulations. This review discusses the underlying molecular mechanism of various nanoformulations of curcumin for the treatment of different cancers. We hope that this study will make a road map for preclinical and clinical investigations of cancer and recommend nano curcumin as a drug of choice for cancer therapy.

scholarly journals TRIM27 interacts with Iκbα to promote the growth of human renal cancer cells through regulating the NF-κB pathway

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Chengwu Xiao ◽  
Wei Zhang ◽  
Meimian Hua ◽  
Huan Chen ◽  
Bin Yang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Prognostic Indicator ◽  
The Cancer Genome Atlas ◽  
Mrna Levels ◽  
Loss Of Function ◽  
Protein Levels ◽  
Kaplan Meier ◽  
Cancer Genome Atlas

Abstract Background The tripartite motif (TRIM) family proteins exhibit oncogenic roles in various cancers. The roles of TRIM27, a member of the TRIM super family, in renal cell carcinoma (RCC) remained unexplored. In the current study, we aimed to investigate the clinical impact and roles of TRIM27 in the development of RCC. Methods The mRNA levels of TRIM27 and Kaplan–Meier survival of RCC were analyzed from The Cancer Genome Atlas database. Real-time PCR and Western blotting were used to measure the mRNA and protein levels of TRIM27 both in vivo and in vitro. siRNA and TRIM27 were exogenously overexpressed in RCC cell lines to manipulate TRIM27 expression. Results We discovered that TRIM27 was elevated in RCC patients, and the expression of TRIM27 was closely correlated with poor prognosis. The loss of function and gain of function results illustrated that TRIM27 promotes cell proliferation and inhibits apoptosis in RCC cell lines. Furthermore, TRIM27 expression was positively associated with NF-κB expression in patients with RCC. Blocking the activity of NF-κB attenuated the TRIM27-mediated enhancement of proliferation and inhibition of apoptosis. TRIM27 directly interacted with Iκbα, an inhibitor of NF-κB, to promote its ubiquitination, and the inhibitory effects of TRIM27 on Iκbα led to NF-κB activation. Conclusions Our results suggest that TRIM27 exhibits an oncogenic role in RCC by regulating NF-κB signaling. TRIM27 serves as a specific prognostic indicator for RCC, and strategies targeting the suppression of TRIM27 function may shed light on future therapeutic approaches.

scholarly journals MBRS-48. IDENTIFICATION OF NOVEL THERAPEUTIC APPROACHES FOR MYC-DRIVEN MEDULLOBLASTOMA

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii406-iii406
Author(s):  
Kübra Taban ◽  
David Pauck ◽  
Mara Maue ◽  
Viktoria Marquardt ◽  
Hua Yu ◽  
...  
Keyword(s):  
New Drugs ◽  
Current Therapy ◽  
Cancer Drug ◽  
Mrna Levels ◽  
Cell Models ◽  
Therapeutic Approaches ◽  
Group 3 ◽  
Novel Therapeutic

Abstract Medulloblastoma (MB) is the most common malignant brain tumor in children and is frequently metastatic at diagnosis. Treatment with surgery, radiation and multi-agent chemotherapy may leave survivors of these brain tumors with long-term deficits as a consequence. One of the four consensus molecular subgroups of MB is the MYC-driven group 3 MB, which is the most malignant type and has a poor prognosis under current therapy. Thus, it is important to discover more effective targeted therapeutic approaches. We conducted a high-throughput drug screening to identify novel compounds showing efficiency in group 3 MB using both clinically established inhibitors (n=196) and clinically-applicable compounds (n=464). More than 20 compounds demonstrated a significantly higher anti-tumoral effect in MYChigh (n=7) compared to MYClow (n=4) MB cell models. Among these compounds, Navitoclax and Clofarabine showed the strongest effect in inducing cell cycle arrest and apoptosis in MYChigh MB models. Furthermore, we show that Navitoclax, an orally bioavailable and blood-brain barrier passing anti-cancer drug, inhibits specifically Bcl-xL proteins. In line, we found a significant correlation between BCL-xL and MYC mRNA levels in 763 primary MB patient samples (Data source: “R2 https://hgserver1.amc.nl”). In addition, Navitoclax and Clofarabine have been tested in cells obtained from MB patient-derived-xenografts, which confirmed their specific efficacy in MYChigh versus MYClow MB. In summary, our approach has identified promising new drugs that significantly reduce cell viability in MYChigh compared to MYClow MB cell models. Our findings point to novel therapeutic vulnerabilities for MB that need to be further validated in vitro and in vivo.

scholarly journals Antagonism of Ultraviolet-Light Mutagenesis by the Methyl-Directed Mismatch-Repair System of Escherichia coli

Genetics ◽  
2000 ◽  
Vol 154 (2) ◽  
pp. 503-512 ◽  
Author(s):  
Hongbo Liu ◽  
Stephen R Hewitt ◽  
John B Hays
Keyword(s):  
Escherichia Coli ◽  
Mismatch Repair ◽  
Excision Repair ◽  
Spontaneous Mutation ◽  
Repair System ◽  
Uv Mutagenesis ◽  
Repair Protein ◽  
Mismatch Repair System

Abstract Previous studies have demonstrated that the Escherichia coli MutHLS mismatch-repair system can process UV-irradiated DNA in vivo and that the human MSH2·MSH6 mismatch-repair protein binds more strongly in vitro to photoproduct/base mismatches than to “matched” photoproducts in DNA. We tested the hypothesis that mismatch repair directed against incorrect bases opposite photoproducts might reduce UV mutagenesis, using two alleles at E. coli lacZ codon 461, which revert, respectively, via CCC → CTC and CTT → CTC transitions. F′ lacZ targets were mated from mut+ donors into mutH, mutL, or mutS recipients, once cells were at substantial densities, to minimize spontaneous mutation prior to irradiation. In umu+ mut+ recipients, a range of UV fluences induced lac+ revertant frequencies of 4–25 × 10−8; these frequencies were consistently 2-fold higher in mutH, mutL, or mutS recipients. Since this effect on mutation frequency was unaltered by an Mfd− defect, it appears not to involve transcription-coupled excision repair. In mut+ umuC122::Tn5 bacteria, UV mutagenesis (at 60 J/m2) was very low, but mutH or mutL or mutS mutations increased reversion of both lacZ alleles roughly 25-fold, to 5–10 × 10−8. Thus, at UV doses too low to induce SOS functions, such as Umu2′D, most incorrect bases opposite occasional photoproducts may be removed by mismatch repair, whereas in heavily irradiated (SOS-induced) cells, mismatch repair may only correct some photoproduct/base mismatches, so UV mutagenesis remains substantial.

Exploring Galleria mellonella larval model to evaluate antibacterial efficacy of Cecropin A (1-7)- Melittin against multi-drug resistant enteroaggregative Escherichia coli

2021 ◽  
Author(s):  
Jess Vergis ◽  
S V S Malik ◽  
Richa Pathak ◽  
Manesh Kumar ◽  
Nitin V Kurkure ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Galleria Mellonella ◽  
In Vivo Model ◽  
Drug Resistant ◽  
Physiological Concentration ◽  
Microbial Drug Resistance ◽  
Cecropin A ◽  
Screening And Identification

Abstract High throughput in vivo laboratory models is need for screening and identification of effective therapeutic agents to overcome microbial drug-resistance. This study was undertaken to evaluate in vivo antimicrobial efficacy of short-chain antimicrobial peptide- Cecropin A (1–7)-Melittin (CAMA) against three multi- drug resistant enteroaggregative Escherichia coli (MDR-EAEC) field isolates in a Galleria mellonella larval model. The minimum inhibitory concentration (MIC; 2.0 mg/L) and minimum bactericidal concentration (MBC; 4.0 mg/L) of CAMA were determined by microdilution assay. CAMA was found to be stable at high temperatures, physiological concentration of cationic salts and proteases; safe with sheep erythrocytes, secondary cell lines and commensal lactobacilli at lower MICs; and exhibited membrane permeabilisation. In vitro time-kill assay revealed concentration- and time-dependent clearance of MDR-EAEC in CAMA-treated groups at 30 min. CAMA- treated G. mellonella larvae exhibited an increased survival rate, reduced MDR-EAEC counts, immunomodulatory effect and proved non-toxic which concurred with histopathological findings. CAMA exhibited either an equal or better efficacy than the tested antibiotic control, meropenem. This study highlights the possibility of G. mellonella larvae as an excellent in vivo model for investigating the host-pathogen interaction, including the efficacy of antimicrobials against MDR-EAEC strains.

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