Investigation the effects of vitreous humor on proliferation and dedifferentiation of differentiated NTERA2 cells / Investigar os efeitos do humor vítreo na proliferação e desdiferenciação de células NTERA2 diferenciadas

Abstract Mammals have a limited capacity to regenerate their tissues and organs. One of the mechanisms associated with natural regeneration is dedifferentiation. Several small molecules such as vitamin C and growth factors could improve reprogramming efficiency. In this study, the NTERA2-D1 (NT2) cells were induced towards differentiation (NT2-RA) with 10-5 M retinoic acid (RA) for three days and then subjected to various amounts of vitreous humor (VH). Results show that the growth rate of these cells was reduced, while this rate was partly restored upon treatment with VH (NT2-RA-VH). Cell cycle analysis with PI method also showed that the numbers of cells at the S phase of the cell cycle in these cells were increased. The levels of SSEA3 and TRA-1-81 antigens in NT2-RA were dropped but they increased in NT2- RA-VH to a level similar to the NT2 cells. The level of SSEA1 had an opposite pattern. Expression of OCT4 gene dropped after RA treatment, but it was recovered in NT2-RA-VH cells. In conclusion, we suggest VH as a potent mixture for improving the cellular reprogramming leading to dedifferentiation.

Resumo Os mamíferos têm uma capacidade limitada de regenerar seus tecidos e órgãos. Um dos mecanismos associados à regeneração natural é a desdiferenciação. Várias moléculas pequenas, como vitamina C e fatores de crescimento, podem melhorar a eficiência da reprogramação. Neste estudo, as células NTERA2-D1 (NT2) foram induzidas à diferenciação (NT2-RA) com ácido retinóico (RA) 10-5 M por três dias e depois submetidas a várias quantidades de humor vítreo (VH). Os resultados mostram que a taxa de crescimento dessas células foi reduzida, enquanto essa taxa foi parcialmente restaurada após o tratamento com VH (NT2-RA-VH). A análise do ciclo celular com o método PI também mostrou que o número de células na fase S do ciclo celular nessas células estava aumentado. Os níveis de antígenos SSEA3 e TRA-1-81 em NT2-RA diminuíram, mas aumentaram em NT2-RA-VH a um nível semelhante ao das células NT2. O nível de SSEA1 teve um padrão oposto. A expressão do gene OCT4 diminuiu após o tratamento com AR, mas foi recuperado em células NT2-RA-VH. Em conclusão, sugerimos o VH como uma mistura potente para melhorar a reprogramação celular levando à desdiferenciação.

Deficiency of two-pore segment channel 2 contributes to systemic lupus erythematosus via regulation of apoptosis and cell cycle / 中华医学杂志(英文版)

BACKGROUND@#Systemic lupus erythematosus (SLE) is a complex autoimmune disease, and the mechanism of SLE is yet to be fully elucidated. The aim of this study was to explore the role of two-pore segment channel 2 (TPCN2) in SLE pathogenesis.@*METHODS@#Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to detect the expression of TPCN2 in SLE. We performed a loss-of-function assay by lentiviral construct in Jurkat and THP-1 cell. Knockdown of TPCN2 were confirmed at the RNA level by qRT-PCR and protein level by Western blotting. Cell Count Kit-8 and flow cytometry were used to analyze the cell proliferation, apoptosis, and cell cycle of TPCN2-deficient cells. In addition, gene expression profile of TPCN2-deficient cells was analyzed by RNA sequencing (RNA-seq).@*RESULTS@#TPCN2 knockdown with short hairpin RNA (shRNA)-mediated lentiviruses inhibited cell proliferation, and induced apoptosis and cell-cycle arrest of G2/M phase in both Jurkat and THP-1 cells. We analyzed the transcriptome of knockdown-TPCN2-Jurkat cells, and screened the differential genes, which were enriched for the G2/M checkpoint, complement, and interleukin-6-Janus kinase-signal transducer and activator of transcription pathways, as well as changes in levels of forkhead box O, phosphatidylinositol 3-kinase/protein kinase B/mechanistic target of rapamycin, and T cell receptor pathways; moreover, TPCN2 significantly influenced cellular processes and biological regulation.@*CONCLUSION@#TPCN2 might be a potential protective factor against SLE.

Rescue and vegetative propagation of Eremanthus erythropappus (DC. ) Ma ish in natural stand / Resgate e propagação vegetativa de Eremanthus erythropappus (DC. ) MacLeish em povoamentos naturais

Abstract The native stands of 'candeia' (Eremanthus erythropappus) have been explored through management plans due to the economic potential of essential oil. The rescue of adult trees, as well as the application of silvicultural techniques that favor the restoration of the stand, can contribute to the genetic conservation of this species. This study's objective was to assess the efficiency of propagation techniques for the rescue of 26 matrices of 'candeia' in a natural managed stand and discussion about the rhizogenesis. In August 2017, trees were induced to regrowth by coppice, followed by exposure and scarification of roots. The emergence of shoots and morphology were evaluated according to the origin (i.e., stump or root). After that period, 19 matrices had their sprouts collected for the preparation of apical cuttings. Indole-3-butyric acid (IBA) was applied at the base of the cuttings. Cutting survival at greenhouse exit (GE), rooting at shade house exit (SHE), morphology and root anatomy were evaluated. In 189 days, the scarification of roots promoted 76.92% of budding. The percentage of sprouted matrices, number of shoots per matrice, length, diameter, and shoot length/diameter ratio increased over time. Only 12.2% of the cuttings survived in GE, and of these, 7.9% rooted in SHE. The cutting resulted in the formation of a clonal mini-garden of 'candeia', with seven of the 19 matrices submitted to propagation. The anatomical analyses showed that bud formation occurs from cell redifferentiation in the phloem parenchyma, and presence of crystals on the walls of the vessel elements of the secondary xylem. The shoots induction from scarification of roots could be used as a silvicultural practice for the reestablishment of the native fragments handle.

Resumo Os povoamentos nativos de candeia (Eremanthus erythropappus) vêm sendo explorados por planos de manejo devido ao potencial econômico do óleo essencial. O resgate de árvores adultas, bem como a aplicação de técnicas silviculturais que favoreçam o restabelecimento do povoamento podem contribuir para a conservação genética dessa espécie. O objetivo desse trabalho foi avaliar a eficiência de técnicas de propagação para o resgate de 26 matrizes de candeia em um povoamento natural manejado e discutir sobre a rizogênese. Em agosto de 2017, as árvores foram induzidas à rebrota por meio da decepa, seguida da exposição e escarificação das raízes. A emissão brotações e morfologia foram avaliadas de acordo com a origem (toco ou raiz). Após esse período, 19 matrizes tiveram as brotações recolhidas para o preparo de estacas apicais, que foram tratadas com ácido indolbutírico (AIB). A sobrevivência das estacas na saída da casa de vegetação (SCV), o enraizamento na saída da casa de sombra (SCS), a morfologia e a anatomia da raiz foram avaliados. Aos 189 dias, a escarificação das raízes resultou em 76,92% de emissão de brotos. O percentual de matrizes brotadas, número de brotos por matriz, comprimento, diâmetro e relação comprimento/diâmetro dos brotos aumentaram ao longo do período avaliado. Somente 12,2% das estacas sobreviveram na SCV e 7,9% enraizaram na SCS. A estaquia resultou na formação de um minijardim clonal de candeia com sete das dezenove matrizes submetidas à propagação. As análises anatômicas mostraram a diferenciação das células na região do parênquima floemático e a presença de cristais de inulina nas paredes dos elementos de vaso do xilema secundário. A indução de brotos radiculares pode ser usada como prática silvicultural visando o restabelecimento de fragmentos nativos manejados.

Effects of Tripterine on Adhesion Molecules and Cell Cycle in Human Acute Promyelocytic Leukemia Model Mice / 中国实验血液学杂志

OBJECTIVE@#To observe the effects of tripterine on adhesion molecules and cell biological characteristics in mice with acute promyelocytic leukemia (APL) tumor.@*METHODS@#Eighteen SCID beige mice were caudal vein injected with NB4 cell lines (5×10@*RESULTS@#The neutrophil decrased and promyelocytes, NB4 cells, B lymphocytes and white blood cells increased in tumor-bearing group as compared with control group (P<0.05), and the expressions of serum P-selectin (P-selectin), soluble vascular adhesion molecule-1 (soluble vascular adhesion molecule-1, sVCAM-1) and soluble intercellular adhesion molecule-1 (soluble intercellular adhesion molecule-1, sICAM-1) all increased (P<0.05). The cell cycle showed that the proportion of G@*CONCLUSION@#Tripterine may not only inhibit the expression of sVCAM-1 and sICAM-1 proteins in APL tumor-bearing mice and reduce the adhesion of tumor cells, but also block tumor cells at G

Crossover patterns under meiotic chromosome program / 亚洲男科学杂志(英文版)

Repairing DNA double-strand breaks (DSBs) with homologous chromosomes as templates is the hallmark of meiosis. The critical outcome of meiotic homologous recombination is crossovers, which ensure faithful chromosome segregation and promote genetic diversity of progenies. Crossover patterns are tightly controlled and exhibit three characteristics: obligatory crossover, crossover interference, and crossover homeostasis. Aberrant crossover patterns are the leading cause of infertility, miscarriage, and congenital disease. Crossover recombination occurs in the context of meiotic chromosomes, and it is tightly integrated with and regulated by meiotic chromosome structure both locally and globally. Meiotic chromosomes are organized in a loop-axis architecture. Diverse evidence shows that chromosome axis length determines crossover frequency. Interestingly, short chromosomes show different crossover patterns compared to long chromosomes. A high frequency of human embryos are aneuploid, primarily derived from female meiosis errors. Dramatically increased aneuploidy in older women is the well-known "maternal age effect." However, a high frequency of aneuploidy also occurs in young women, derived from crossover maturation inefficiency in human females. In addition, frequency of human aneuploidy also shows other age-dependent alterations. Here, current advances in the understanding of these issues are reviewed, regulation of crossover patterns by meiotic chromosomes are discussed, and issues that remain to be investigated are suggested.

Methodology of Differentiation of Bone Marrow Cells into Megakaryocytes in Vitro / 中国实验血液学杂志

OBJECTIVE@#To explore the method for inducing the differentiation of bone marrow cells into megakaryocytes in vitro so as to use for evaluating the activity of traditional Chinese medicines.@*METHODS@#The bone marrow cells were separated from femurs and tibias of mice. The experiments were divided into 4 groups: control (no adding cytokines), TPO (adding 50 ng/ml TPO), TPO+SCF (50 ng/ml+50 ng/ml) and TPO+SCF+IL-6+IL-9 (50 ng/ml+50 ng/ml+20 ng/ml+20 ng/ml). The bone marrow cells in 4 groups were cultured in vitro for 6 d. Then the cell growth status was observed by the inverted microscopy, and the cell count was detected by using the automatic cell counter. The ratio and absolute count of megakaryocytes were detected by flow cytometry.@*RESULTS@#Compared with control, three induction methods could stimulate the differentiation of bone marrow cells into megakaryocytes in vitro. TPO could slightly enhance the differentiation of bone marrow cells into megakaryocytes. Both the combination of TPO and SCF, and the combination of TPO, SCF, IL-6 and IL-9 could intensively stimulate proliferation of bone morrow cells and promote the differentiation of bone marrow cells into megakaryocytes. The addition of IL-6 and IL-9 could decrease the proliferation of non-megakaryocytes, but promote the differentiation of bone marrow cells into megakaryocytes.@*CONCLUSION@#The optimized differentiation of bone marrow cells into megakaryocytes has been completed by co-induction regimen of TPO, SCF, IL-6 and IL-9, which can be used to screen and evaluate traditional Chinese medicines promoting formation of platelets.

Effects of cell division protein-encoding genes knockout on solvent formation and cell morphology in Clostridium acetobutylicum / 生物工程学报

Clostridium acetobutylicum is an important strain for bio-butanol formation. In recent years, gene-editing technology is widely used for developing the hyper-butanol-production strains. In this study, three genes (cac1251, cac2118 and cac2125) encoding cell division proteins (RodA, DivIVA and DivIB) in C. acetobutylicum were knocked out. The cac2118-knockout strain had changed its cell morphology to spherical-shape during the solventogenesis, and obtained a higher butanol yield of 0.19 g/g, increasing by 5.5%, compared with the wild type strain. The glucose utilization and butanol production of cac1251-knockout strain decreased by 33.9% and 56.3%, compared the with wild type strain, reaching to 47.3 g/L and 5.6 g/L. The cac1251-knockout strain and cac2125-knockout strain exhibited poor cell growth with cell optical density decreased by 40.4% and 38.3%, respectively, compared with that of the wild type strain. The results indicate that cell division protein DivIVA made the differences in the regulation of cell morphology and size. Cell division proteins RodA and DivIB played significant roles in the regulation of cell division, and affected cell growth, as well as solventogenesis metabolism.

El caseinato de sodio y la caseína α inhiben la proliferación de la línea celular mieloide de ratón 32D clone 3 (32Dcl3) mediante el TNF-α / Sodium caseinate and alfa-casein inhibit proliferation of the mouse myeloid cell line 32D clone 3 (32Dcl3) via TNF-α

Resumen Introducción. Se ha demostrado que el caseinato de sodio y sus componentes (caseínas α, β y κ) inhiben la proliferación de la línea celular hematopoyética de ratón 32D clone 3 (32Dcl3) e inducen su diferenciación hacia macrófagos. Se sabe que la caseína α induce la producción de IL-1β y que esta última citocina inhibe la proliferación celular mediante la producción del factor de necrosis tumoral alfa (TNF-α), pero se desconoce si el caseinato de sodio y las caseínas inducen la producción de TNF y si este es el responsable de la inhibición de la proliferación. Objetivo. Evaluar si el caseinato de sodio y las caseínas α, β y κ inhiben la proliferación de la línea celular 32Dcl3 mediante la producción de TNF-α. Materiales y métodos. Se usaron diferentes concentraciones de caseinato de sodio y de las caseínas α, β y κ en las células 32Dcl3. Posteriormente, se evaluaron la viabilidad celular mediante una prueba con el MTT [3-(4,5-dimetiltiazol-2-ilo)-2,5-difeniltetrazol], la inducción de apoptosis con la citometría de flujo y la síntesis del TNF-α con el ELISA. Además, se hicieron pruebas de neutralización con anti-TNF-α en células 32Dcl3 tratadas con caseinato de sodio y caseína α, y se evaluó la proliferación celular. Resultados. Se encontró que el caseinato de sodio y las caseínas α, β y κ reducían la proliferación de la línea celular 32Dcl3 sin afectar la viabilidad, y que solo el caseinato y la caseína α inducían la apoptosis y la liberación al medio de TNF-α. La proliferación de células 32Dcl3 tratadas con caseinato y caseína α se restableció al usar anticuerpos anti-TNF-α. Conclusión. El TNF-α fue el principal responsable de la inhibición de la proliferación en las células 32Dcl3 tratadas con caseinato de sodio o caseína α.

Abstract Introduction: Sodium caseinate (CS) and its components (alpha-casein, beta-casein, and kappa-casein) have been shown to inhibit the proliferation of the mouse hematopoietic 32D clone 3 (32Dcl3) cell line and induce its differentiation into macrophages. It is well-known that alpha-casein induces IL-1β production and that this cytokine inhibits the proliferation via the production of tumor necrosis factor alpha (TNF-alpha), but it is not known if CS and the caseins inhibit the proliferation via TNF-alpha production. Objective: To evaluate if CS and alpha-casein, beta-casein and kappa-casein inhibit the proliferation on 32Dcl3 cell line via TNF-alpha. Materials and methods: We used different concentrations of CS, alpha-casein, beta-casein and kappa-casein in 32Dcl3 cells to evaluate cell proliferation. We assessed cell viability by MTT, induction to apoptosis by flow cytometry, and TNF-alpha synthesis by ELISA. Additionally, we performed anti-TNF-alpha neutralization assays on 32Dcl3 cells treated with CS and alpha-casein and we evaluated proliferation. Results: The results showed that CS, alpha-casein, beta-casein, and kappa-casein reduced proliferation of the 32Dcl3 cell line without affecting the viability and that only CS and alpha-casein induced apoptosis and the release of TNF-alpha. The 32Dcl3 cells treated with CS and alpha-casein reestablished their proliferation by using anti-TNF-alpha antibodies. Conclusion: TNF-alpha was the main responsible for the inhibition of proliferation in 32Dcl3 cells treated with CS or alpha-casein.

Cell Division Cycle Associated 8 Is a Key Regulator of Tamoxifen Resistance in Breast Cancer / 한국유방암학회지

PURPOSE: Breast cancer (BC) is one of the most common malignancies globally, and millions of women worldwide are diagnosed with BC every year. Up to 70% of BC patients are estrogen receptor (ER)-positive. Numerous studies have shown that tamoxifen has a significant therapeutic effect on both primary and metastatic ER-positive BC patients. Although tamoxifen is currently one of the most successful therapeutic agents for BC, a significant proportion of patients will eventually become resistant to tamoxifen, leading to tumor recurrence and metastasis. Knowledge about the development of tamoxifen resistance in BC patients is still limited. METHODS: We applied a loss-and-gain method to study the biological functional role of cell division cycle associated 8 (CDCA8) in tamoxifen resistance in BC cells. RESULTS: We found that CDCA8 was significantly elevated in tamoxifen-resistant BC cells. Knockdown of CDCA8 expression significantly inhibited the proliferation of tamoxifen-resistant BC cells and reduced their resistance to tamoxifen. In contrast, overexpression of CDCA8 promoted the growth of tamoxifen-sensitive BC cells and induced their resistance to tamoxifen. CONCLUSION: In this study, we reported that CDCA8 is a key regulator of tamoxifen resistance in BC, suggesting that CDCA8 may serve as a potential therapeutic target for BC treatment.

Diverse Effects of Small Molecule Inhibitors on Actin Cytoskeleton Dynamics in HIV-1 Infection

The dynamics of the actin cytoskeleton plays a pivotal role in the process of cell division, the transportation of organelles, vesicle trafficking and cell movement. Human immunodeficiency virus type 1 (HIV-1) hijacks the actin dynamics network during the viral entry and migration of the pre-integration complex (PIC) into the nucleus. Actin dynamics linked to HIV-1 has emerged as a potent therapeutic target against HIV infection. Although some inhibitors have been intensely analyzed with regard to HIV-1 infection, their effects are sometimes disputed and the exact mechanisms for actin dynamics in HIV infection have not been well elucidated. In this study, the small molecules regulating HIV-1 infection from diverse inhibitors of the actin dynamic network were screened. Two compounds, including Chaetoglobosin A and CK-548, were observed to specifically bar the viral infection, while the cytochalasin family, 187-1, N-WASP inhibitor, Rho GTPase family inhibitors (EHop-016, CID44216842, and ML-141) and LIMK inhibitor (LIM domain kinase inhibitor) increased the viral infection without cytotoxicity within a range of ~ µM. However, previously known inhibitory compounds of HIV-1 infection, such as Latrunculin A, Jasplakinolide, Wiskostatin and Swinholide A, exhibited either an inhibitory effect on HIV-1 infection combined with severe cytotoxicity or showed no effects. Our data indicate that Chaetoglobosin A and CK-548 have considerable potential for development as new therapeutic drugs for the treatment of HIV infection. In addition, the newly identified roles of Cytochalasins and some inhibitors of Rho GTPase and LIMK may provide fundamental knowledge for understanding the complicated actin dynamic pathway when infected by HIV-1. Remarkably, the newly defined action modes of the inhibitors may be helpful in developing potent anti-HIV drugs that target the actin network, which are required for HIV infection.

Telomere Biology in Mood Disorders: An Updated, Comprehensive Review of the Literature

Major psychiatric disorders are linked to early mortality and patients afflicted with these ailments demonstrate an increased risk of developing physical diseases that are characteristically seen in the elderly. Psychiatric conditions like major depressive disorder, bipolar disorder and schizophrenia may be associated with accelerated cellular aging, indicated by shortened leukocyte telomere length (LTL), which could underlie this connection. Telomere shortening occurs with repeated cell division and is reflective of a cell’s mitotic history. It is also influenced by cumulative exposure to inflammation and oxidative stress as well as the availability of telomerase, the telomere-lengthening enzyme. Precariously short telomeres can cause cells to undergo senescence, apoptosis or genomic instability; shorter LTL correlates with compromised general health and foretells mortality. Important data specify that LTL may be reduced in principal psychiatric illnesses, possibly in proportion to exposure to the ailment. Telomerase, as measured in peripheral blood monocytes, has been less well characterized in psychiatric illnesses, but a role in mood disorder has been suggested by preclinical and clinical studies. In this manuscript, the most recent studies on LTL and telomerase activity in mood disorders are comprehensively reviewed, potential mediators are discussed, and future directions are suggested. An enhanced comprehension of cellular aging in psychiatric illnesses could lead to their re-conceptualizing as systemic ailments with manifestations both inside and outside the brain. At the same time this paradigm shift could identify new treatment targets, helpful in bringing about lasting cures to innumerable sufferers across the globe.

Expression of polo-like kinase 1 in pre-implantation stage murine somatic cell nuclear transfer embryos

Somatic cell nuclear transfer (SCNT) has various applications in research, as well as in the medical field and animal husbandry. However, the efficiency of SCNT is low and the accurate mechanism of SCNT in murine embryo development is unreported. In general, the developmental rate of SCNT murine embryos is lower than in vivo counterparts. In previous studies, polo-like kinase 1 (Plk1) was reported to be a crucial element in cell division including centrosome maturation, cytokinesis, and spindle formation. In an initial series of experiments in this study, BI2536, a Plk1 inhibitor, was treated to in vivo-fertilized embryos and the embryos failed to develop beyond the 2-cell stage. This confirmed previous findings that Plk1 is crucial for the first mitotic division of murine embryos. Next, we investigated Plk1's localization and intensity by immunofluorescence analysis. In contrast to normally developed embryos, SCNT murine embryos that failed to develop exhibited two types of Plk1 expressions; a low Plk1 expression pattern and ectopic expression of Plk1. The results show that Plk1 has a critical role in SCNT murine embryos. In conclusion, this study demonstrated that the SCNT murine embryos fail to develop beyond the 2-cell stage, and the embryos show abnormal Plk1 expression patterns, which may one of the main causes of developmental failure of early SCNT murine embryos.

Antitumor profiles and cardiac electrophysiological effects of aurora kinase inhibitor ZM447439

Aurora kinases inhibitors, including ZM447439 (ZM), which suppress cell division, have attracted a great deal of attention as potential novel anti-cancer drugs. Several recent studies have confirmed the anti-cancer effects of ZM in various cancer cell lines. However, there have been no studies regarding the cardiac safety of this agent. We performed several cytotoxicity, invasion and migration assays to examine the anti-cancer effects of ZM. To evaluate the potential effects of ZM on cardiac repolarisation, whole-cell patch-clamp experiments were performed with human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and cells with heterogeneous cardiac ion channel expression. We also conducted a contractility assay with rat ventricular myocytes to determine the effects of ZM on myocardial contraction and/or relaxation. In tests to determine in vitro efficacy, ZM inhibited the proliferation of A549, H1299 (lung cancer), MCF-7 (breast cancer) and HepG2 (hepatoma) cell lines with IC₅₀ in the submicromolar range, and attenuated the invasive and metastatic capacity of A549 cells. In cardiac toxicity testing, ZM did not significantly affect I(Na), I(Ks) or I(K1), but decreased I(hERG) in a dose-dependent manner (IC₅₀: 6.53 µM). In action potential (AP) assay using hiPSC-CMs, ZM did not induce any changes in AP parameters up to 3 µM, but it at 10 µM induced prolongation of AP duration. In summary, ZM showed potent broad-spectrum anti-tumor activity, but relatively low levels of cardiac side effects compared to the effective doses to tumor. Therefore, ZM has a potential to be a candidate as an anti-cancer with low cardiac toxicity.

Genetic features associated with ¹⁸F-FDG uptake in intrahepatic cholangiocarcinoma

PURPOSE: In intrahepatic cholangiocarcinoma (iCCA), genetic characteristics on ¹⁸F-fluorodeoxyglucose (¹⁸F-FDG)-PET scans are not yet clarified. If specific genetic characteristics were found to be related to FDG uptake in iCCA, we can predict molecular features based on the FDG uptake patterns and to distinguish different types of treatments. In this purpose, we analyzed RNA sequencing in iCCA patients to evaluate gene expression signatures associated with FDG uptake patterns. METHODS: We performed RNA sequencing of 22 cases iCCA who underwent preoperative ¹⁸F-FDG-PET, and analyzed the clinical and molecular features according to the maximum standard uptake value (SUVmax). Genes and biological pathway which are associated with SUVmax were analyzed. RESULTS: Patients with SUVmax higher than 9.0 (n = 9) had poorer disease-free survival than those with lower SUVmax (n = 13, P = 0.035). Genes related to glycolysis and gluconeogenesis, phosphorylation and cell cycle were significantly correlated with SUVmax (r ≥ 0.5). RRM2, which is related to the toxicity of Gemcitabine was positively correlated with SUVmax, and SLC27A2 which is associated with Cisplastin response was negatively correlated with SUVmax. According to the pathway analysis, cell cycle, cell division, hypoxia, inflammatory, and metabolism-related pathways were enriched in high SUVmax patients. CONCLUSION: The genomic features of gene expression and pathways can be predicted by FDG uptake features in iCCA. Patients with high FDG uptake have enriched cell cycle, metabolism and hypoxic pathways, which may lead to a more rational targeted treatment approach.

Cytotoxic and genotoxic potential of Ginkgo biloba L, in industrialized and without-additive forms / Potencial citotóxico e genotóxico de Ginkgo biloba L, nas formas industrializada e não aditivada

The toxic potential at the cellular level of industrialized Ginkgo biloba L. leaves was evaluated in meristematic cells of Allium cepa at concentrations of 0.1; 0.2 and 0.4 mg/ml. The industrialized products, from four pharmaceutical laboratories, were identified as A, B, C and D. Cell-level toxicity of dehydrated ginkgo leaf tea was also evaluated at concentrations of 0.15; 0.30 and 0.60 mg/ml. Dehydrated products were purchased from herbalists certified by ANVISA. The roots were exposed to teas and processed products for 24 and 48 hours. The results were submitted to the Chi-square test at 5%. However, industrialized ginkgo products at all concentrations caused antiproliferative effect. Also, the products purchased in pharmacies did not induce significant changes to root meristems. Therefore, industrialized ginkgo promoted cytotoxicity, however, they were not genotoxic to the bioassay used.

Avaliou-se, em células meristemáticas de raízes de Allium cepa, o potencial tóxico em nível celular de folhas de Ginkgo biloba L. industrializadas, nas concentrações 0,1; 0,2 e 0,4 mg/mL. Os produtos industrializados, oriundos de quatro laboratórios farmacêuticos, foram identificados como A, B, C e D. Também avaliou-se a toxicidade em nível celular de chás de folhas de ginkgo desidratadas, nas concentrações 0,15; 0,30 e 0,60 mg/mL. Os produtos desidratados foram adquiridos em ervanários certificados pela ANVISA. As raízes ficaram expostas aos chás e produtos industrializados por 24 e 48 horas. Os resultados obtidos foram submetidos ao teste Qui-quadrado, a 5%. No entanto, os produtos de ginkgo industrializados, em todas as concentrações, causaram efeito antiproliferativo. Ainda, os produto adquiridos em farmácias não induziram alterações em número significativo aos meristemas de raízes. Portanto, os ginkgos industrializados promoveram citotoxicidade, porém, não foram genotóxicos frente ao bioensaio utilizado.

Effect of danusertib on cell cycle, apoptosis and autophagy of hepatocellular carcinoma HepG2 cells / 南方医科大学学报

OBJECTIVE@#To investigate the effect of danusertib (Danu), an inhibitor of Aurora kinase, on the proliferation, cell cycle, apoptosis, and autophagy of hepatocellular carcinoma HepG2 cells and explore the underlying mechanisms.@*METHODS@#MTT assay was used to examine the effect of Danu on the viability of HepG2 cells to determine the IC50 of Danu. The effect of Danu on cell cycle distribution, apoptosis and autophagy were determined using flow cytometry. Western blotting was used to detect the expressions of the proteins related to cell cycle, apoptosis and autophagy. Chloroquine was used to suppress Danuinduced autophagy to test the apoptosis-inducing effect of Danu.@*RESULTS@#Danu significantly inhibited the proliferation of HepG2 cells with IC of 39.4 μmol and 14.4 μmol at 24 h and 48 h, respectively. Danu caused cell cycle arrest in G/M phase in HepG2 cells and led to polyploidy accumulation via up-regulating the expressions of p53 and p21 and down-regulating the expressions of cyclin B1 and DC2. Danu also caused apoptosis of HepG2 cells through up-regulating the expressions of Bax, Puma, cleaved caspase-3, cleaved caspase-9, cleaved PARP and cytochrome C and down-regulating the expressions of Bcl-xl and Bcl-2. Danu induced autophagy via activating AMPK signaling and inhibiting PI3K/PTEN/AKT/mTOR axis, and inhibition of Danu-induced autophagy with chloroquine enhanced the pro-apoptotic effect of Danu.@*CONCLUSIONS@#Danu inhibits cell proliferation and induces cell cycle arrest in G/M phase, apoptosis and cytoprotective autophagy in HepG2 cells.

Can Tissue Cilia Lengths and Urine Cilia Proteins Be Markers of Kidney Diseases? / 전남의대학술지

The primary cilium is an organelle which consists of a microtubule in the core and a surrounding cilia membrane, and has long been recognized as a “vestigial organelle”. However, new evidence demonstrates that the primary cilium has a notable effect on signal transduction in the cell and is associated with some genetic and non-genetic diseases. In the kidney, the primary cilium protrudes into the Bowman's space and the tubular lumen from the apical side of epithelial cells. The length of primary cilia is dynamically altered during the normal cell cycle, being shortened by retraction into the cell body at the entry of cell division and elongated at differentiation. Furthermore, the length of primary cilia is also dynamically changed in the cells, as a result and/or cause, during the progression of various kidney diseases including acute kidney injury and chronic kidney disease. Notably, recent data has demonstrated that the shortening of the primary cilium in the cell is associated with fragmentation, apart from retraction into the cell body, in the progression of diseases and that the fragmented primary cilia are released into the urine. This data reveals that the alteration of primary cilia length could be related to the progression of diseases. This review will consider if primary cilia length alteration is associated with the progression of kidney diseases and if the length of tissue primary cilia and the presence or increase of cilia proteins in the urine is indicative of kidney diseases.

Development of a Novel Nonradioisotopic Assay and Cdc25B Overexpression Cell Lines for Use in Screening for Cdc25B Inhibitors

PURPOSE: The cyclin-dependent kinase 1 (Cdk1) and cyclin B complex performs important roles in the transition from the G2 to M phase in the cell cycle through removal of inhibitory phosphates on Cdk1, and Cdc25B, which is a dual-specific phosphatase, mediates these dephosphorylation events. However, measuring Cdc25B activity by existing methods is hampered by inadequate nonspecific substrates and the need to use a radiolabeled isotope. The present study aimed to develop an improved method with which to properly measure Cdc25B activity using a novel nonradioisotopic assay and Cdc25B overexpression cell lines. MATERIALS AND METHODS: A nonradioisotopic Cdk1 kinase assay, based on Western blotting for retinoblastoma protein and histone H1, was used to analyze Cdc25B activity. Also, stable Cdc25B2 and Cdc25B3 overexpression HeLa cell lines were constructed using the tetracycline-regulated expression system and were applied as a tool for screening for inhibitors of Cdc25B. RESULTS: The present study developed and optimized a nonradioisotopic assay method to properly measure Cdc25B activity. Furthermore, we constructed stable Cdc25B2 and Cdc25B3 overexpression HeLa cell lines for the establishment of a strong assay system with which to evaluate the specificity of Cdc25B inhibitors under conditions similar to the intracellular environment. These methods were confirmed as useful tools for measuring Cdc25B activity. CONCLUSION: The nonradioisotopic Cdk1 kinase assay and Cdc25B overexpression cell lines developed in this study can be conveniently used as tools for screening inhibitors of Cdc25B phosphatase as anticancer drugs.

Single-cell transcriptomics reveals gene signatures and alterations associated with aging in distinct neural stem/progenitor cell subpopulations

Aging associated cognitive decline has been linked to dampened neural stem/progenitor cells (NSC/NPCs) activities manifested by decreased proliferation, reduced propensity to produce neurons, and increased differentiation into astrocytes. While gene transcription changes objectively reveal molecular alterations of cells undergoing various biological processes, the search for molecular mechanisms underlying aging of NSC/NPCs has been confronted by the enormous heterogeneity in cellular compositions of the brain and the complex cellular microenvironment where NSC/NPCs reside. Moreover, brain NSC/NPCs themselves are not a homogenous population, making it even more difficult to uncover NSC/NPC sub-type specific aging mechanisms. Here, using both population-based and single cell transcriptome analyses of young and aged mouse forebrain ependymal and subependymal regions and comprehensive "big-data" processing, we report that NSC/NPCs reside in a rather inflammatory environment in aged brain, which likely contributes to the differentiation bias towards astrocytes versus neurons. Moreover, single cell transcriptome analyses revealed that different aged NSC/NPC subpopulations, while all have reduced cell proliferation, use different gene transcription programs to regulate age-dependent decline in cell cycle. Interestingly, changes in cell proliferation capacity are not influenced by inflammatory cytokines, but likely result from cell intrinsic mechanisms. The Erk/Mapk pathway appears to be critically involved in regulating age-dependent changes in the capacity for NSC/NPCs to undergo clonal expansion. Together this study is the first example of using population and single cell based transcriptome analyses to unveil the molecular interplay between different NSC/NPCs and their microenvironment in the context of the aging brain.

Chinese medicinal compound CFF-1 induces the apoptosis and cycle-arrest of prostate cancer cells via the PI3K/AKT/FOXO1 signaling pathway / 中华男科学杂志

Objective@#To explore the apoptosis-inducing effect of the Chinese medicinal compound CFF-1 on prostate cancer cells and its related molecular mechanisms.@*METHODS@#Normal prostate WPMY-1 cells and prostate cancer LNCaP, CWR22Rv1, PC3 and DU145 cells were treated in dehydrated alcohol with CFF-1 at 0, 2, 5, or 10 mg/ml for 24 hours. Then the viability of the prostate cells was detected by morphological observation, MTT and CCK-8 assay, nuclear condensation and disruption measured by DAPI staining, the cell cycle and apoptosis calculated by flow cytometry, the activity of the PI3K/AKT/FOXO1 signaling pathway and the expressions of its downstream apoptosis- and cycle-related proteins determined by Western blot.@*RESULTS@#CFF-1 significantly arrested the cell cycle in the G1 phase, decreased the cell viability and increased the nuclear condensation and disruption in a dose-dependent manner, and elevated the apoptosis rate of prostate cancer cells. At the molecular level, CFF-1 dose-dependently reduced the activity of the PI3K/AKT signaling pathway and phosphorylation of the FOXO1 protein, increased the transcription activity of FOXO1, and eventually regulated the expressions of cell apoptosis- and cycle-related genes.@*CONCLUSIONS@#The Chinese medicinal compound CFF-1 can significantly inhibit the growth, arrest the cycle, and induce the apoptosis of prostate cancer cells by decreasing the activity of the PI3K/AKT/FOXO1 signaling pathway, which suggests its potential clinical application value in the treatment of prostate cancer.