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INTRODUCTION: This study delved into the role of Kinase Insert Domain Receptor (KDR) and its associated miRNAs in renal cell carcinoma through an extensive computational analysis. The potential of our findings to guide future research in this area is significant. METHODS: Our methods, which included the use of UALCAN and GEPIA2 databases, as well as miRDB, MirDIP, miRNet v2.0, miRTargetLink, MiEAA v2.1, TarBase v8.0, INTERNET, and miRTarBass, were instrumental in identifying the regulation of miRNA associated with KDR expression. The predicted miRNA was validated with the TCGA-KIRC patients' samples by implementing CancerMIRNome. The TargetScanHuman v8.0 was implemented to identify the associations between human miRNAs and KDR. A Patch Dock server analyzed the interactions between hsa-miR-200b-3p-KDR and hsa-miR-200b-3p with KDR. RESULTS: The KDR expression rate was investigated in the Kidney Renal Cell Carcinoma (KIRC) samples, and adjacent normal tissues revealed that the expression rate was significantly higher than the normal samples, which was evident from the strong statistical significance (P = 1.63e-12). Likely, the KDR ex-pression rate was estimated as high at tumor grade 1 and gradually decreased till the metastasis grade, reducing the survival rate of the KIRC patients. To identify these signals early, we predicted a miRNA that could trigger the expression of KDR. Furthermore, we uncovered the potential associations between miR-200c-3p expressions by regulating KDR towards the progression of KIRC. DISCUSSION: Upon examining the outcome, it became evident that miR-200c-3p was significantly down-regulated in KIRC compared to the normal samples. Moreover, the negative correlation was obtained for hsa-miR-200c-3p (R = - 0.276) along with the KDR expression describing that the increased rate of hsa-miR-200c-3p might reduce the KDR expression rate, which may suppress the KIRC initiation or progres-sion. CONCLUSION: The in-silico analysis indicated that the significant increase in KDR expression during the initiation of KIRC could serve as an early diagnostic marker. Moreover, KDR could be utilized to identify advancements in KIRC stages. Additionally, hsa-miR-200c-3p was identified as a potential regulator capable of downregulating and upregulating KDR expression among the 24 miRNAs screened. This find-ing holds promise for future research endeavors. Concurrent administration of the FDA-approved 5-fluor-ouracil with KIRC drugs, such as sorafenib, zidovudine, and everolimus, may have the potential to en-hance the therapeutic efficacy in downregulating hsa-miR-200c-3p. However, further in vitro studies are imperative to validate these findings and gain a comprehensive understanding of the intricate regulatory interplay involving hsa-miR-200c-3p, KDR, 5-fluorouracil, and other FDA-approved drugs for the treat-ment of KIRC. This will facilitate the identification of KIRC stage progression and its underlying pre-ventative mechanisms.
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Expression of the constitutively active serine/threonine kinase Akt in oligodendrocytes results in enhanced myelination in the CNS. Here, we have examined the effects of this Akt overexpression on optic nerve structure and on optic nerve function, assessed using the visual evoked potential (VEP). Transgenic mice have been generated with the Plp promoter driving expression of a modified form of Akt, in which aspartic acids are substituted for Thr308 and Ser473. These Plp-Akt-DD (Akt-DD) mice, and littermate controls, were studied at different ages. Optic nerves were examined anatomically at 2 and 6 months of age. At 2 months of age, optic nerves were substantially thicker in Akt-DD mice, reflecting an increase in myelination of optic nerve axons. By electron microscopy, myelin thickness was increased in Akt-DD optic nerve, with extended paranodal domains having excess paranodal loops, and the density of nodes of Ranvier was reduced, relative to control mice. We recorded VEPs in response to strobe flash ganzfeld stimuli presented after overnight dark- and light-adapted conditions at ages ranging from 1 to 10 months. It was possible to record a clear VEP from Akt-DD mice at all ages examined. At 1 month of age, VEP implicit times were somewhat shorter in Akt-DD transgenic mice than in control animals. Beyond 6months of age, VEP latencies were consistently delayed in Akt-DD transgenic mice. These abnormalities did not reflect an alteration in retinal function as there were no significant differences between ERGs obtained from control or Akt-DD transgenic mice. In young mice, the somewhat faster responses may reflect improved transmission due to increased myelination of optic nerve axons. In older mice, where the Akt-DD optic nerve is markedly thicker than control, it is remarkable that optic nerves continue to function.
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Fibras Nerviosas Mielínicas/patología , Nervio Óptico/patología , Trastornos de la Visión/patología , Animales , Electrorretinografía/métodos , Potenciales Evocados Visuales/fisiología , Ratones , Ratones Transgénicos , Vaina de Mielina/enzimología , Vaina de Mielina/patología , Fibras Nerviosas Mielínicas/enzimología , Proteína Oncogénica v-akt/biosíntesis , Nervio Óptico/enzimología , Estimulación Luminosa/métodos , Tiempo de Reacción/fisiología , Trastornos de la Visión/enzimologíaRESUMEN
Mammalian target of rapamycin (mTOR), a well known Akt substrate, regulates multiple cellular functions including cell growth and protein synthesis. The current study identifies a novel role of the Akt/mTOR pathway as a regulator of CNS myelination. Previously, we showed that overexpressing constitutively active Akt in oligodendrocytes in a transgenic mouse model induces enhanced CNS myelination, with no changes in the proliferation or survival of oligodendrocyte progenitor or mature cells. The present study focused on the signaling mechanisms regulating this hypermyelination induced by Akt. Activation of mTOR and its downstream substrates (p70S6 kinase and S6 ribosomal protein) was observed in Akt-overexpressing oligodendrocytes. When mTOR signaling was inhibited chronically in vivo with rapamycin starting at 6 weeks of age, the observed hypermyelination was reduced to approximately the amount of myelin seen in wild-type mice. mTOR inhibition had little impact on wild-type myelination between 6 and 12 weeks of age, suggesting that, in normal adults, myelination is relatively complete and is no longer regulated by mTOR signaling. However, when mTOR was chronically inhibited in young adult wild-type mice, myelination was reduced. These results suggest that, during active myelination, the major Akt signal regulating CNS myelination is the mTOR pathway.
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Sistema Nervioso Central/metabolismo , Proteínas de la Mielina/metabolismo , Proteína Oncogénica v-akt/metabolismo , Proteínas Quinasas/metabolismo , Transducción de Señal/fisiología , 2',3'-Nucleótido Cíclico Fosfodiesterasas/metabolismo , Animales , Animales Recién Nacidos , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Sistema Nervioso Central/efectos de los fármacos , Sistema Nervioso Central/ultraestructura , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/fisiología , Inmunosupresores/farmacología , Ratones , Ratones Transgénicos , Microscopía Electrónica de Transmisión/métodos , Proteínas de la Mielina/genética , Proteína Proteolipídica de la Mielina/genética , Vaina de Mielina/efectos de los fármacos , Vaina de Mielina/metabolismo , Fibras Nerviosas Mielínicas/efectos de los fármacos , Fibras Nerviosas Mielínicas/metabolismo , Fibras Nerviosas Mielínicas/ultraestructura , Proteínas del Tejido Nervioso/metabolismo , Factor de Transcripción 2 de los Oligodendrocitos , Oligodendroglía/efectos de los fármacos , Oligodendroglía/metabolismo , Proteína Oncogénica v-akt/genética , Transducción de Señal/efectos de los fármacos , Sirolimus/farmacología , Serina-Treonina Quinasas TOR , Factores de TiempoRESUMEN
The serine/threonine kinase Akt regulates multiple cellular functions. The current studies identify a new role for Akt in CNS myelination. In earlier studies on cultured oligodendrocytes, we showed that neuregulin signals through phosphatidylinositol-3'-OH kinase and Akt to enhance survival of oligodendrocytes. However, when transgenic animals were generated that overexpressed constitutively active Akt in oligodendrocytes and their progenitor cells, no enhanced survival of oligodendrocytes or progenitors was found. No alteration in the proliferation or death of progenitors was noted. In contrast, the major impact of Akt overexpression in oligodendrocytes was enhanced myelination. Most interestingly, oligodendrocytes in these mice continued actively myelinating throughout life. Thus, expression of constitutively active Akt in oligodendrocytes and their progenitor cells generated no more oligodendrocytes, but dramatically more myelin. The increased myelination continued as these mice aged, resulting in enlarged optic nerves and white matter areas. In older animals with enlarged white matter areas, the density of oligodendrocytes was reduced, but because of the increased area, the total number of oligodendrocytes remained comparable with wild-type controls. Interestingly, in these animals, overexpression of Akt in Schwann cells did not impact myelination. Thus, in vivo, constitutively active Akt enhances CNS myelination but not PNS myelination and has no impact developmentally on oligodendrocyte number. Understanding the unique aspects of Akt signal transduction in oligodendrocytes that lead to myelination rather than uncontrolled proliferation of oligodendrocyte progenitor cells may have important implications for understanding remyelination in the adult nervous system.
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Sistema Nervioso Central/fisiología , Regulación de la Expresión Génica/fisiología , Vaina de Mielina/fisiología , Proteína Oncogénica v-akt/fisiología , Factores de Edad , Animales , Bromodesoxiuridina/metabolismo , Muerte Celular/fisiología , Proliferación Celular , Sistema Nervioso Central/citología , Proteínas Fluorescentes Verdes/biosíntesis , Ratones , Ratones Transgénicos , Microscopía Electrónica de Transmisión/métodos , Proteína Proteolipídica de la Mielina/genética , Vaina de Mielina/ultraestructura , Oligodendroglía/fisiología , Oligodendroglía/ultraestructura , Proteína Oncogénica v-akt/genética , Nervio Óptico/fisiología , Nervio Óptico/ultraestructura , Nervio Ciático/fisiología , Nervio Ciático/ultraestructura , Serina/metabolismo , Treonina/metabolismoRESUMEN
Virtual screening emerged as an important tool in our quest to access novel drug like compounds. There are a wide range of comparable and contrasting methodological protocols available in screening databases for the lead compounds. The number of methods and software packages which employ the target and ligand based virtual screening are increasing at a rapid pace. However, the general understanding on the applicability and limitations of these methodologies is not emerging as fast as the developments of various methods. Therefore, it is extremely important to compare and contrast various protocols with practical examples to gauge the strength and applicability of various methods. The review provides a comprehensive appraisal on several of the available virtual screening methods to-date. Recent developments of the docking and similarity based methods have been discussed besides the descriptor selection and pharmacophore based searching. The review touches upon the application of statistical, graph theory based methods machine learning tools in virtual screening and combinatorial library design. Finally, several case studies are undertaken where the virtual screening technology has been applied successfully. A critical analysis of these case studies provides a good platform to estimate the applicability of various virtual screening methods in the new lead identification and optimization.