RESUMO
The antitumor activity of resveratrol, a polyphenolic compound found mainly in grapes, has been studied in several types of cancer. In bladder cancer, its antiproliferative effects have already been demonstrated; however, its mechanism of action is not completely understood. The aim of this study was to evaluate resveratrol antitumor activity (12.5, 25, 50, 100, 150, 200, and 250 µM) and its possible mechanisms of action in bladder tumor cells with different TP53 gene status (RT4, grade 1, TP53 wild type; 5637-grade 2 and T24-grade 3, TP53 mutated). Cell proliferation, clonogenic survival, morphological changes, cell cycle progression, apoptosis rates, genotoxicity, global methylation, immunocytochemistry for p53 and PCNA and relative expression profiles of the AKT, mTOR, RASSF1A, HOXB3, SRC, PLK1, and DNMT1 were evaluated. Resveratrol decreased cell proliferation and induced DNA damage in all cell lines. Regarding the long-term effects, resveratrol reduced the number of colonies in all cell lines; however, TP53 wild type cells were more resistant. Increased rates of apoptosis were found in the TP53 wild type cells and this was accompanied by AKT, mTOR, and SRC downregulation. In addition, the resveratrol antiproliferative effects in wild type TP53 cells were accompanied by modulation of the DNMT1 gene. In the TP53 mutated cells, cell cycle arrest at S phase with PLK1 downregulation was observed. Additionally, there was modulation of the HOXB3/RASSF1A pathway and nuclear PCNA reduction in the highest-grade cells. In conclusion, resveratrol has antiproliferative activity in bladder tumor cells; however, the mechanisms of action are dependent on TP53 status. Environ. Mol. Mutagen., 60:740-751, 2019. © 2019 Wiley Periodicals, Inc.
Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Resveratrol/farmacologia , Proteína Supressora de Tumor p53/genética , Neoplasias da Bexiga Urinária/tratamento farmacológico , Apoptose/efeitos dos fármacos , Apoptose/genética , Proteínas de Ciclo Celular/biossíntese , Linhagem Celular Tumoral , Proliferação de Células/genética , Dano ao DNA/genética , Humanos , Antígeno Nuclear de Célula em Proliferação/biossíntese , Proteínas Serina-Treonina Quinases/biossíntese , Proteínas Proto-Oncogênicas/biossíntese , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase S do Ciclo Celular/genética , Proteína Supressora de Tumor p53/biossíntese , Quinase 1 Polo-LikeRESUMO
BACKGROUND: The objective was to assess histopathological changes and the expression of proliferating cell nuclear antigen (PCNA), Bcl-2, suppressor of cytokine signaling (SOCS) 1 and 3, Vimentin, TWIST1, and Cdh 1 and 2 in early stages of experimental oral carcinogenesis process using a shorter period of exposure to 4-nitroquinoline oxide (4-NQO) model. METHODS: In this study, 20 rats were divided into control group (n = 10), sacrificed on the first day of the experiment, and experimental group (n = 10) treated with 50 ppm of 4-NQO solution dissolved in drinking water for 8 and 12 weeks. The histological sections were stained with H&E or subjected to immunohistochemistry for detecting PCNA, Bcl-2, SOCS 1 and 3, and STAT 3. Some specimens were used for verification of Vimentin expression, Cdh 1, Cdh 2, and TWIST1 by RT-qPCR. RESULTS: At both 8 and 12 weeks, morphological changes occurred mainly in the posterior portion of the tongue and were limited to the epithelial tissue, including moderate to severe dysplasia at 8 weeks, and severe dysplasia with exacerbation of atypical cells at 12 weeks. Expression of SOCS 1 and 3 increased from 8 to 12 weeks (P < 0.05), whereas STAT 3 expression was reduced mainly at 12 weeks (P < 0.05) in comparison with the control group. The expression of all epithelial-mesenchymal transition markers (EMT) was increased after 12 weeks, reaching statistical significance (P < 0.05) for Cdh 1 and 2. CONCLUSIONS: Together, the results suggested that overexpression of Bcl-2, SOCS 1 and 3, and Cdh 1 and 2 is associated with the early neoplasic changes in modified 4-nitroquinoline 1-oxide-induced murine oral cancer model.
Assuntos
4-Nitroquinolina-1-Óxido , Biomarcadores Tumorais/biossíntese , Carcinógenos , Neoplasias Bucais/induzido quimicamente , Neoplasias Bucais/metabolismo , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Caderinas/biossíntese , Caderinas/genética , Modelos Animais de Doenças , Transição Epitelial-Mesenquimal , Masculino , Neoplasias Bucais/genética , Neoplasias Bucais/patologia , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Antígeno Nuclear de Célula em Proliferação/biossíntese , Antígeno Nuclear de Célula em Proliferação/genética , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Proteínas Proto-Oncogênicas c-bcl-2/genética , Ratos , Proteína 1 Supressora da Sinalização de Citocina/biossíntese , Proteína 1 Supressora da Sinalização de Citocina/genética , Proteína 3 Supressora da Sinalização de Citocinas/biossíntese , Proteína 3 Supressora da Sinalização de Citocinas/genética , Proteína 1 Relacionada a Twist/biossíntese , Proteína 1 Relacionada a Twist/genética , Vimentina/biossíntese , Vimentina/genéticaRESUMO
The purpose of our study was to observe the effects of luteolin on the expression of the genes ICAM-1, LFA-3, and PCNA in H22 hepatoma tissue. Sixty ICR (Institute of Cancer Research) mice with H22 hepatoma were randomly divided into five groups: a normal saline control group, low-, medium-, and high-dose luteolin groups, and a cyclophosphamide group. The mice were euthanized the day after administration withdrawal and subcutaneous tumor tissue was extracted. Quantitative fluorescence RT-PCR was used to detect the expression of ICAM-1, LFA-3, and PCNA in H22 hepatoma tissue in the mice. Luteolin was found to up-regulate the expression of ICAM-1 in H22 hepatoma tissue, of which the middle-dose group had the most obvious effect, showing a significant difference (P < 0.01) as compared to the normal saline group. Each dose group of luteolin significantly down-regulated the expression of LFA-3 in H22 hepatoma tissue, showing significant differences as compared to the saline control group (P < 0.01). The medium- and high-dose luteolin groups significantly reduced the expression of PCNA in H22 hepatoma tissue of ICR mice, where the effect of the high-dose group was the most obvious, and the difference between the two luteolin groups and the normal saline group was statistically significant (P < 0.01). Luteolin may inhibit tumor angiogenesis and tumor cell proliferation by down-regulation of LFA- 3 and PCNA and up-regulation of ICAM-1 in tumor tissue of tumor-bearing mice, thereby achieving its anti-tumor effect.
Assuntos
Antígenos CD58/biossíntese , Carcinoma Hepatocelular/genética , Molécula 1 de Adesão Intercelular/biossíntese , Neoplasias Hepáticas/genética , Neovascularização Patológica/genética , Antígeno Nuclear de Célula em Proliferação/biossíntese , Animais , Antígenos CD58/genética , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Fluoruracila/administração & dosagem , Regulação Neoplásica da Expressão Gênica , Molécula 1 de Adesão Intercelular/genética , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , Luteolina/administração & dosagem , Camundongos , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/patologia , Antígeno Nuclear de Célula em Proliferação/genéticaRESUMO
BACKGROUND: The therapeutic potential of adult stem cells in the treatment of chronic diseases is becoming increasingly evident. In the present study, we sought to assess whether treatment with mesenchymal stem cells (MSCs) efficiently retards progression of chronic renal failure (CRF) when administered to experimental models of less severe CRF. METHODS: We used two renal mass reduction models to simulate different stages of CRF (5/6 or 2/3 mass renal reduction). Renal functional parameters measured were serum creatinine (SCr), creatinine clearance (CCr), rate of decline in CCr (RCCr), and 24-h proteinuria (PT24h). We also evaluated renal morphology by histology and immunohistochemistry. MSCs were obtained from bone marrow aspirates and injected into the renal parenchyma of the remnant kidneys of both groups of rats with CRF (MSC5/6 or MSC2/3). RESULTS: Animals from groups MSC5/6 and CRF2/3 seemed to benefit from MSC therapy because they showed significantly reduction in SCr and PT24h, increase in CCr and slowed the RCCr after 90 days. Treatment reduced glomerulosclerosis but significant improvement did occur in the tubulointerstitial compartment with much less fibrosis and atrophy. MSC therapy reduced inflammation by decreasing macrophage accumulation proliferative activity (PCNA-positive cells) and fibrosis (α-SM-actin). Comparisons of renal functional and morphological parameters responses between the two groups showed that rats MSC2/3 were more responsive to MSC therapy than MSC5/6. CONCLUSION: This study showed that MSC therapy is efficient to retard CRF progression and might be more effective when administered during less severe stages of CRF.
Assuntos
Falência Renal Crônica/terapia , Transplante de Células-Tronco Mesenquimais/métodos , Actinas/biossíntese , Actinas/genética , Animais , Proliferação de Células , Creatinina/metabolismo , Progressão da Doença , Feminino , Fibrose/patologia , Glomerulosclerose Segmentar e Focal/patologia , Glomerulosclerose Segmentar e Focal/terapia , Rim/patologia , Falência Renal Crônica/patologia , Testes de Função Renal , Macrófagos/patologia , Células-Tronco Mesenquimais , Antígeno Nuclear de Célula em Proliferação/biossíntese , Antígeno Nuclear de Célula em Proliferação/genética , Proteinúria/metabolismo , Ratos , Ratos WistarRESUMO
BACKGROUND: The aim of this study is to compare antimicrobial photodynamic therapy (aPDT) as an adjunctive therapy to scaling and root planing (SRP) for the treatment of experimentally induced periodontitis in rats with ovariectomy (OVX) that are or are not treated with estrogen replacement. METHODS: A total of 270 female rats were divided into three groups: 1) normal rats; 2) rats with OVX; and 3) rats with OVX with estrogen replacement. Periodontal disease was induced through the introduction of a cotton thread around the mandibular left first molar. After 7 days, the ligature was removed, and the rats were randomly divided into the following treatment groups: 1) SRP plus saline solution; 2) SRP plus low-level laser therapy (LLLT); and 3) SRP plus toluidine blue O irrigation followed by LLLT. Ten rats from each group were euthanized at days 7, 15, and 30 after dental treatment. Bone loss (BL) in the furcation region was evaluated using histometric and immunohistochemical analyses. RESULTS: aPDT treatment resulted in reduced BL compared with SRP treatment at all time points. Additionally, rats treated with aPDT exhibited reduced numbers of tartrate-resistant acid-phosphatase-positive cells and more proliferating cell nuclear antigen-positive cells in all treatment groups regardless of estrogen status. Whereas rats treated with aPDT showed weak immunoreactivity to the receptor activator of nuclear factor-κ B ligand at day 7 post-treatment, strong osteoprotegerin immunoreactivity was observed at day 15 post-treatment. CONCLUSION: aPDT is an effective adjunctive therapy for the treatment of periodontitis in rats with OVX that are or are not given estrogen replacement therapy.
Assuntos
Perda do Osso Alveolar/tratamento farmacológico , Terapia de Reposição de Estrogênios , Periodontite/tratamento farmacológico , Fotoquimioterapia , Ligante RANK/antagonistas & inibidores , Perda do Osso Alveolar/microbiologia , Animais , Quimioterapia Adjuvante , Raspagem Dentária , Feminino , Interleucina-1/metabolismo , Interleucina-6/metabolismo , Terapia com Luz de Baixa Intensidade , Ovariectomia , Periodontite/microbiologia , Antígeno Nuclear de Célula em Proliferação/biossíntese , Ratos , Fator de Necrose Tumoral alfa/metabolismoRESUMO
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⻹) from Days 0 to 6 and 500 ng mL⻹ from Days 7 to 12) increased follicular diameter and resulted in increased antrum formation (P<0.05). Alone, 200 ng mL⻹ 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⻹ bovine serum albumin, 10 µg mL⻹ insulin, 5.5 µg mL⻹ transferrin, 5 ng mL⻹ selenium, 2 mM glutamine, 2mM hypoxanthine and 50 µg mL⻹ ascorbic acid (α-MEMâº). Comparisons of uncultured (0.2 mm) and α-MEM⺠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.
Assuntos
Hormônio Foliculoestimulante/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Fator 9 de Diferenciação de Crescimento/metabolismo , Técnicas de Maturação in Vitro de Oócitos/veterinária , Oogênese , Folículo Ovariano/metabolismo , RNA Mensageiro/metabolismo , Matadouros , Animais , Bovinos , Sobrevivência Celular , Feminino , Líquido Folicular/enzimologia , Líquido Folicular/metabolismo , Glucuronosiltransferase/antagonistas & inibidores , Glucuronosiltransferase/biossíntese , Glucuronosiltransferase/genética , Glucuronosiltransferase/metabolismo , Hialuronan Sintases , Isoenzimas/antagonistas & inibidores , Isoenzimas/biossíntese , Isoenzimas/genética , Isoenzimas/metabolismo , Oócitos/citologia , Oócitos/enzimologia , Oócitos/metabolismo , Folículo Ovariano/citologia , Folículo Ovariano/crescimento & desenvolvimento , Antígeno Nuclear de Célula em Proliferação/biossíntese , Antígeno Nuclear de Célula em Proliferação/química , Antígeno Nuclear de Célula em Proliferação/genética , Antígeno Nuclear de Célula em Proliferação/metabolismo , Proteoglicanas/antagonistas & inibidores , Proteoglicanas/biossíntese , Proteoglicanas/genética , Proteoglicanas/metabolismo , Receptores do FSH/antagonistas & inibidores , Receptores do FSH/biossíntese , Receptores do FSH/genética , Receptores do FSH/metabolismo , Técnicas de Cultura de Tecidos/veterináriaRESUMO
Proliferating cell nuclear antigen (PCNA), a member of the sliding clamp family of proteins, interacts specifically with DNA replication and repair proteins through a small peptide motif called the PCNA-interacting protein or PIP box. PCNA is recognized as one of the key proteins involved in DNA metabolism. In the present study, the recombinant PCNA from Litopenaeus vannamei (LvPCNA) was heterologously overexpressed and purified using metal ion-affinity chromatography. Crystals suitable for diffraction grew overnight using the hanging-drop vapour-diffusion method. LvPCNA crystals belong to space group C2 with unit-cell parameters a=144.6, b=83.4, c=74.3â Å, ß=117.6°. One data set was processed to 3â Å resolution, with an overall Rmeas of 0.09 and a completeness of 93.3%. Initial phases were obtained by molecular replacement using a homology model of LvPCNA as the search model. Refinement and structural analysis are underway. This report is the first successful crystallographic analysis of a marine crustacean decapod shrimp (L. vannamei) proliferating cell nuclear antigen.
Assuntos
Proteínas de Artrópodes/química , Penaeidae , Antígeno Nuclear de Célula em Proliferação/química , Animais , Proteínas de Artrópodes/biossíntese , Clonagem Molecular , Cristalização , Cristalografia por Raios X , Escherichia coli , Antígeno Nuclear de Célula em Proliferação/biossínteseRESUMO
BACKGROUND: Kinins are released during dermal injury and inflammation and seem to contribute to the pathogenesis of cutaneous diseases. OBJECTIVE: Participation of kinins in skin inflammatory process was evaluated using knockout mice and non-peptide kinin receptor antagonists. METHODS: Chronic skin inflammation was induced by multiple applications of TPA in mice ear. RESULTS: The B(2) knockout mice (B(2)(-/-)) showed a significant increase of ear weight (23 ± 10%) and epidermal cellular hyperproliferation and acanthosis formation upon histological analysis when compared with wildtype mice. Also, evaluation of PCNA levels by Western blot and immunohistochemistry confirmed the increase in the epidermis hyperproliferation in the ear skin of B(2)(-/-) mice. In contrast, no modification in these parameters was detected in B(1) knockout mice (B(1)(-/-)). However, mice lacking both kinin receptors (B(1)B(2)(-/-)) presented a considerable reduction of epidermis thickness and in PCNA levels. Following the establishment of skin inflammation (5th day of TPA application) treatment with the non-peptide antagonists SSR 240612 (B(1) receptor antagonist), FR 173657 (B(2) receptor antagonist), or SSR 240612 plus FR 173657 topically applied, caused a significant inhibition of ear weight (20 ± 5%, 34 ± 4% and 32 ± 6%, respectively). In the histological analysis, the antagonists produced a reduction in epidermal hyperplasia and acanthosis formation; but the treatment with a combination of the two antagonists did not increase efficacy. CONCLUSION: Kinin receptors seem to be involved in the control of the keratinocyte hyperproliferative process, and non-peptide kinin receptor antagonists may be useful tools in the treatment of hyperproliferative skin disorders.
Assuntos
Receptor B1 da Bradicinina/metabolismo , Receptor B2 da Bradicinina/metabolismo , Dermatopatias/patologia , Administração Tópica , Animais , Proliferação de Células , Dioxóis/farmacologia , Feminino , Inflamação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Antígeno Nuclear de Célula em Proliferação/biossíntese , Quinolinas/farmacologia , Sulfonamidas/farmacologia , Acetato de Tetradecanoilforbol/farmacologiaRESUMO
OBJECTIVE: To evaluate the effect of melatonin both on the ovaries of pinealectomized female rats through histomorphometric analysis and on steroid receptors, proliferating cell nuclear antigen (PCNA), and vascular endothelial growth factor (VEGF) expression. DESIGN: Experimental study. SETTING: Federal University of São Paulo, Brazil. ANIMAL(S): Forty female rats. INTERVENTION(S): Forty rats were divided equally into four groups: GI-vehicle without surgery; GII--surgery without removal of the pineal gland (sham); GIII--pinealectomized with vehicle; and GIV--pinealectomized with melatonin treatment. After treatment for 3 consecutive months, the animals were killed and their ovaries removed for analysis. MAIN OUTCOME MEASURE(S): Estrogen and progesterone receptors, histologic and immunohistochemical analysis. RESULT(S): The GIII samples presented signals of proliferation on ovarian surface epithelium and interstitial cells as well as high expressions of PCNA and VEGF in those structures compared with GI, GII, and GIV. Also, the levels of progesterone receptor (fmol/g) in ovaries of GIII (250.6 ± 32.4) were significantly lower than in those of GI (429.0 ± 23,8), GII (442.3 ± 30.2), and GIV (564.1 ± 78.7). The levels of progesterone in GIII were superior to those in GI, GII, and GIV. CONCLUSION(S): Our findings suggest that melatonin may attenuate proliferation in ovarian structures and increase the number of luteal bodies as well as the levels of progesterone receptor.
Assuntos
Melatonina/fisiologia , Ovário/metabolismo , Glândula Pineal/cirurgia , Antígeno Nuclear de Célula em Proliferação/biossíntese , Receptores de Esteroides/biossíntese , Fator A de Crescimento do Endotélio Vascular/biossíntese , Animais , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Regulação da Expressão Gênica , Ovário/citologia , Glândula Pineal/metabolismo , Ligação Proteica/genética , Ratos , Ratos Wistar , Receptores de Esteroides/genética , Transdução de Sinais/genética , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
OBJECTIVE: to analyze the immunohistochemical expression of P27 and CD34 markers as prognostic factors in patients with localized prostate cancer. METHODS: analysis of 100 patients with localized prostate cancer submitted to curative surgery. We carried out the usual histological preparation, followed by immunohistochemistry to detect the accumulation of P27 and CD34 protein followed by statistical analysis. RESULTS: in the evaluation of P27 marker and on the correlation with the variables we found significant difference in Gleason score with positive expression (positive P27) related to lower mean PSA (p = 0.091), lower Gleason score (p < 0.0001) and smaller tumor area in CD34 (p = 0.036). Regarding the CD34 marker at the tumor area, it was observed that the smaller the positive CD34, the lower the PSA value (p < 0.0001) and lower the Gleason score (r = 0.5726, p < 0.0001), and the higher the positive CD34, the higher the staging (r = 0.3305, p <0.0001) and the chance of recurrence (p = 0.002). Patients with higher stage also displayed larger positive CD34 areas (p < 0.0001). CONCLUSION: the markers CD34 and P27 are associated with events specific to prostate cancer, however, only CD34 was able to determine the possibility of biochemical recurrence.