Hepatic protective effect of grape seed proanthocyanidin extract against Gleevec-induced apoptosis, liver Injury and Ki67 alterations in rats

ABSTRACT Gleevec (imatinib) is an antineoplastic chemotherapeutic agent used in the treatment of many types of cancer. The current study was conducted to examine the possible modifying effects of grape seeds proanthocyandins extract (GSPE) against apoptosis, liver injury and Ki67 alterations induced by Gleevec in male rats. 40 male albino rats were equally divided into four groups (First and second groups were control and GSPE groups; third group was Gleevec group and fourth group was treated with Gleevec and GSPE). Gleevec induced elevations in P53 and depletion of Bcl2 levels in liver tissues were compared with the control group. Liver sections in rats treated with Gleevec exhibited marked cellular infiltrations, vacuolar degeneration hepatocytes, numerous apoptotic cells, and congestion in central and portal veins, as well as a significant increase in the proliferating of Ki67 after Gleevec injection as compared with control group. In contrast, treatment with Gleevec and GSPE showed a moderate to good degree of improvement in hepatocytes with a significant increase in Ki67, a decrease in P53 and an increase in Bcl2 levels in liver tissues compared to treatment with Gleevec. Therefore, Gleevec induces apoptosis, injury and Ki67 changes in rat liver, whereas GSPE modulates these alternations.

Evaluation of claspin as a proliferation marker in human cancer and normal tissues.

Claspin is a nuclear protein involved in DNA replication and the DNA damage response. Its structural and functional properties suggest that it may represent a potentially useful proliferation marker. To this end, a monoclonal antibody was generated and the expression of claspin was investigated in normal fibroblasts and various cancer cell lines, as well as in tumour and normal tissues from patients with primary epithelial carcinomas. Immunoblotting analysis confirmed the specificity of the antibody, while immunohistochemistry demonstrated its applicability in archival material. In normal cells and tissues, claspin expression was weak, whereas increased levels were observed in cancer cell lines and tumour specimens. Claspin staining correlated strongly with Ki67 staining in both normal (p < 0.001) and tumour tissues (p < 0.001). However, the labelling index (LI) of claspin was consistently lower than that of Ki67, suggesting that claspin expression may be limited to a narrower part of the cell cycle. Co-localization assays with cyclin A and cell synchronization experiments indicated that claspin expression coincides with the S phase. Interestingly, the relative increase of the claspin LI in tumour samples compared with normal tissues was significantly higher (14-fold) than that of the Ki67 LI (five-fold), suggesting that claspin may be a more sensitive marker of aberrant proliferation.

The role of 5-HT1A receptors in the proliferation and survival of progenitor cells in the dentate gyrus of the adult hippocampus and their regulation by corticoids.

These experiments explore the role of 5-HT1A receptors in the regulation of cell proliferation in the dentate gyrus of the intact and adrenalectomized adult rat. Depleting 5-HT with p-chlorophenylalanine (300 mg/kg initially followed by 100 mg/kg/day) or stimulating 5-HT1A receptors with 8-OH-DPAT (1 mg/kg or 2 mg/kg, s.c. injections twice daily) for 14 days had no effect on cell proliferation as measured by Ki-67 or BrdU (5-bromo-3-deoxyuridine) immunocytochemistry in the dentate gyrus. However, combined treatment with p-chlorophenylalanine followed by 8-OH-DPAT significantly increased cell proliferation compared with p-chlorophenylalanine alone. Micro-injection of the 5-HT neurotoxin 5,7-dihydroxytryptamine into the fimbria-fornix (3.0 microg/side) and the cingulate bundle (1.8 microg/side) depleted hippocampal 5-HT locally but did not change cell proliferation 3 weeks after the surgery. However, 8-OH-DPAT (1 mg/kg, twice daily) stimulated cell proliferation in the dentate gyrus of hippocampal 5-HT-depleted rats compared with controls. These results suggest that 5-HT(1A) modulates cell proliferation in the hippocampus by a direct post-synaptic effect. Previous studies demonstrate that adrenalectomy increases hippocampal 5-HT1A receptor expression and binding, and thus we investigated whether the effect of adrenalectomy on cell proliferation and survival was dependent on the activity of the 5-HT1A receptors. In contrast to the null effect following twice-daily s.c. injection, 8-OH-DPAT (2.0 mg/kg/day) delivered by s.c. osmotic pumps increased proliferation in intact rats. The 5-HT1A antagonist WAY-100635 (1.5 mg/kg/day also delivered by osmotic pump) by itself did not alter cell proliferation, confirming that reduced serotonin activity does not change proliferation, but blocked the effect of 8-OH-DPAT. However, WAY-100635 could not block the stimulating action of adrenalectomy cell proliferation. 5-HT1A mRNA expression was not altered in the hippocampus by adrenalectomy. Thus, the effect of adrenalectomy on cell proliferation and survival is not 5-HT1A dependent, despite the interaction between 5-HT1A and corticosterone.

Valor pronóstico de los factores clinicopatológicos Ki67, ciclina D1, ciclina D3 y CDK4 en el carcinoma gástrico / Pronostic value of clinicopathologic factors Ki67, cyclin D1, cyclin D3 and CDK4 in gastric carcinoma

- Propósito: Conocer el valor pronóstico de la ciclina Dl, ciclina D3, cdk4 y Ki67, estudiados por métodos inmunohistoquímicos, junto con las características clinicopatológicos de los carcinomas gástricos.- Métodos y resultados: Realizamos estudio inmunohistoquimicos de material incluido en parafina para ciclina D1, ciclina D3, cdk4 y Ki67 en 74 pacientes con carcinoma gástrico. Las inmunotinciones para ciclinas D1, D3 y cdk4 así el índice de proliferación de Ki67, el grado histológico y el tipo histológico (según la clasificación de Lauren) se compararon con la supervivencia. El 97 por ciento de los casos eran Ki67 positivos, el 29 por ciento para ciclina D1, el 23 por ciento para ciclina D3 y el 35 por ciento para cdk4. El análisis multivariante sólo mostró correlación entre el Ki67 (PI) (p<0,01) y la supervivencia. En el análisis univariante el grado histológico también se correlaciona con la supervivencia (p<0,03). La expresión de ciclina D3 se relaciona con cdk4 (p<0,001) y Ki67 (PI) (p<0,02) y la expresión de ciclina D1 con el grado histológico (p<0,03).- Conclusiones: Nuestros resultados sugieren que un índice de proliferación elevado de Ki67 y el grado histológico son marcadores de mal pronóstico. La sobreexpresión de la ciclina D1 podría tener un importante papel en la proliferación celular. La relación entre ciclina D3, cdk4 y Ki67 podrían explicarse por su papel a lo largo del ciclo celular. (AU)

Proliferation marker pKi-67 affects the cell cycle in a self-regulated manner.

The proliferation marker pKi-67 is commonly used in research and pathology to detect proliferating cells. In a previous work, we found the protein to be associated with regulators of the cell cycle, controlling S-phase progression, as well as entry into and exit from mitosis. Here we investigate whether pKi-67 has a regulative effect on the cell cycle itself. For that purpose we cloned four fragments of pKi-67, together representing nearly the whole protein, and an N-terminal pKi-67 antisense oligonucleotide into a tetracycline inducible gene expression system. The sense fragments were C-terminally modified by addition of either a nuclear localization sequence (NLS) or a STOP codon to address the impact of their intracellular distribution. FACS based cell cycle analysis revealed that expression of nearly all pKi-67 domains and the antisense oligonucleotide led to a decreased amount of cells in S-phase and an increased number of cells in G(2)/M- and G(1)-phase. Subsequent analysis of the endogenous pKi-67 mRNA and protein levels revealed that the constructs with the most significant impact on the cell cycle were able to silence pKi-67 transcription as well. We conclude from the data that pKi-67 influences progression of S-phase and mitosis in a self-regulated manner and, therefore, effects the cell cycle checkpoints within both phases. Furthermore, we found pKi-67 mediates an anti-apoptotic effect on the cell and we verified that this marker, although it is a potential ribosomal catalyst, is not expressed in differentiated tissues with a high transcriptional activity.

Proliferation of rhesus ovarian surface epithelial cells in culture: lack of mitogenic response to steroid or gonadotropic hormones.

Ovarian cancer is the most lethal gynecological cancer, and approximately 90% of ovarian cancers derive from the ovarian surface epithelium (OSE), yet the biology of the OSE is poorly understood. Factors associated with increased risk of non-hereditary ovarian cancer include the formation of inclusion cysts, effects of reproductive hormones and the number of ovulations experienced in a woman's lifetime. Distinguishing between these factors is difficult in vivo, but cultured OSE cells are viable tools for some avenues of research. Here we establish rhesus macaque OSE cultures and demonstrate that these cells express cytokeratin, vimentin, N-cadherin, ER-alpha, and PR but are negative for E-cadherin. We show that these cells activate MAPK and proliferate in response to extracellular calcium, as do human and rat OSE. In contrast, the gonadotropic hormones FSH (4-400 IU/liter), LH (8.5-850 IU/liter), and human CG (10-1000 IU/liter) fail to stimulate proliferation. We find that concentrations of progesterone and estrogen normally present in follicles just before ovulation ( approximately 1000 ng/ml) significantly decrease the number of mitotically active rhesus macaque OSE cells as determined by PCNA labeling, total cell count, and (3)H-thymidine uptake, whereas lower steroid concentrations have no effect.