Effects of ß-glucan from S. cerevisiae on the expression of Casp9, Ccna2 and Sod1 genes in MCF-7 cells

ß-Glucans (ßG) are polysaccharides widely distributed in nature with chemopreventive properties. The aim of this study was to investigate the effects of ßG and the combined treatment with doxorubicin (Dox) on cell viability and mRNA levels of genes involved in cell cycle, apoptosis and antioxidant response. ßG was not cytotoxic. The mRNA levels of CCNA2of cells exposed to ß-glucan was upregulated and the exposure to Dox decreased the expression, while the combination led to an upregulation. Modulation of mRNA levels of CASP9suggest that ßG could inhibit promotion and progression steps of carcinogenesis, eliminatingneoplastic cells. The upregulation of CCNA2gene in combined treatment could be occurred due to ability of ßG in restoring the cell cycle distribution pattern after treatment with Dox. The upregulation of SOD1suggests that ßG can enhance the intracellular antioxidant defense, reducing the levels of superoxide dismutase induced by Dox. This response could reduce oxidative damage and attenuate tissue damage during chemotherapeutic treatment. Our data suggest that the drug combination may be less effective in killing tumor cells than the treatment with Dox alone. Thus, future studies should carefully consider this effect on indication of ßG during chemotherapy.Keywords:caspase-9; cyclin A2; superoxide dismutase 1; cell cycle; antioxidant.Received on July 2, 2020.Accepted on February 7, 2022.IntroductionGlucans are polysaccharides widely distributed in nature and oftenstudied due to chemopreventive properties. They are constituent of the cell wall of plants (oats and barley), algae, bacteria and fungi. ß-glucans (ßG)have a common structure comprising a main chain of ß-(1,3) and/or ß-(1,4) D-glucopyranosyl unit and they differ in length and branching structures. ßG of Saccharomyces cerevisiaehave 1→6 side branches while those of bacteria have 1→4 side branches (Chan, Chan, & Sze, 2009). ßGcan prevent DNA damage induced by chemical and physical agents (Ghavami,Goliaei, Taghizadeh, & Nikoofar, 2014). Some authors showed its significant efficacy in preventing mutagenic effects caused by doxorubicin, cyclophosphamide and cisplatin (Tohamy, El-Ghor, El-Nahas, & Noshy, 2003), methyl methanesulfonate (Oliveira et al., 2007)and hydrogen peroxide (Slamenová, 2003). Moreover, some studies have related the antioxidant ability of ßGagainst reactive free radicals formed by endogenous metabolic processes or exogenous chemicals (Tsiapali et al., 2001; Slamenová,2003; Sener, Eksioglu-Demiralp, Cetiner, Ercan, & Yegen, 2006; Guerra Dore et al., 2007; Kofuji et al., 2012; Lei et al., 2015). Yeast-derived ßGhave modulating action of humoral and cellular immune responses (Vetvicka et al., 2007).This activity provides protection to the organism against infections and cancer development (Samuelsen, Schrezenmeir, & Knutsen, 2014; Roudbary, Daneshmand, Hajimorad, Roudbarmohammadip, & Hassan, 2015). Despite postulated modes of action by which ß-glucan works are lacking information about the molecular mechanisms involved in the chemopreventive activity of this polysaccharide. In addition, compounds with chemopreventive properties can contribute to reduce side effects and toxicity during the chemotherapeutic treatment. Therefore, the aim of this study was to investigate the effects of ßG and the combined treatment with doxorubicin (Dox) on the expression of genes related with apoptosis (CASP9), cell cycle control (CCNA2)and antioxidant defense (SOD1)in human breast cancer MCF-7 cells. Doxorubicin (Dox) was chosen because it is one of the most used chemotherapeutic agent for cancer treatment. The limitation on the use of Dox in cancer treatment is the lack of selectivity against cancer cells and, consequently, its toxicity to patients.(AU)

CCNA1 gene as a potential diagnostic marker in papillary thyroid cancer.

The malignancy that most affects the endocrine system is thyroid neoplasm, with an increasing incidence over the years. The most prevalent histological type of the carcinomas that affect the thyroid gland is papillary carcinoma with a prevalence of 80 % worldwide. The current diagnostic methodology may present inconclusive results, emphasizing the need for new effective and sensitive techniques to aid the diagnosis. For this, it is necessary to understand molecular and protein mechanisms in the identification of diagnostic and predictive markers in the lesions. The Cyclin A1 protein, encoded by the CCNA1 gene, is an important cell cycle regulator, belonging to the MAPK/ERK signaling pathway directly involved with thyroid cancer. The aim of this study was to evaluate the CCNA1 gene and Cyclin A1 protein expression in papillary thyroid carcinoma, follicular thyroid carcinoma, and benign thyroid lesions, by real time quantitative PCR and immunohistochemistry analysis, respectively, to verify their roles as potential diagnostic and predictive markers to future applications in the clinical routine. Overexpression of CCNA1 gene was observed in the papillary carcinoma group compared to the normal group (P = 0.0023), benign lesions (P = 0.0011), colloid goiter (P = 0.0124), and follicular carcinoma (P = 0.0063). No differential expression was observed in the papillary primary tumor group from negative lymph nodes compared with the one from positive lymph nodes (P = 0.3818). Although an increased expression of Cyclin A1 was observed in the PTC group compared to the other one in the IHC analysis, no significant difference was observed (Fisher's exact Test). A Cyclin A1 overexpression was detected with weak to mid-moderate immunoreactivity in the benign group (k = 0.56), (score 1.5); mid-moderate to moderate in the goiter group (k = 0.58); weak in the FTC group (k = 0.33); and mid-moderate to moderate in the PTC group (k = 0.48). Due to the small sample size in the IHC analysis and to the fact that not all RNA is translated into protein, the diagnostic potential of Cyclin A1 could not be assessed. However, these findings highlight the potential of the CCNA1 gene as a diagnostic marker for papillary thyroid carcinoma.

Cyclin A in nonfunctioning pituitary adenomas.

PURPOSE: Assess cyclin A in nonfunctioning pituitary adenomas (NFPA) and compare its expression in non-invasive and non-proliferative tumors with invasive and proliferative tumors (12× higher risk of recurrence). METHODS: Quantitative real time polymerase chain reaction to analyze cyclin A using normal pituitary gland as reference. Fold change (FC) > 1 was considered as increased. Tumor invasion was based on Knosp criteria (grades 3-4 considered invasive) and proliferation on the presence of at least two of three criteria: Ki-67 ≥ 3%; mitoses > 2/10; positive p53. Both groups were compared with Mann-Whitney test considering p value < 0.05 as statistically significant. RESULTS: Thirty-one patients with NFPA were included. Tumors were mainly of gonadotrophic origin (74.2%), followed by corticotrophic (19.4%) and lactotrophic (3.2%) origins and null-cell adenomas (3.2%). Median tumor diameter was 3.5 cm (1.8-8.0) and Ki-67 was 3.0% (0.3-11%). Sixteen patients had tumors classified as non-invasive and non-proliferative and 15 as invasive and proliferative. Median FC was 0.31 in all tumors (0.13-1.94). Cyclin A was not related to invasion or proliferation (FC 0.41 in non-invasive and non-proliferative tumors and FC 0.30 in invasive and proliferative tumors; p = 0.968). Four (12.9%) patients had tumors that exhibited increased cyclin A [median FC of 1.04 (1.02-1.94)]-three of gonadotrophic origin and one null-cell adenoma, with two tumors classified as non-invasive and non-proliferative and two tumors classified as invasive and proliferative. Median tumor diameter in these samples was 3.4 cm (2.4-3.6) and Ki-67 was 5.1% (2-11%). CONCLUSIONS: Cyclin A was increased in a minority of NFPA and does not seem to be related to invasion or proliferation.

The integration of HR-HPV increases the expression of cyclins A and E in cytologies with and without low-grade lesions.

BACKGROUND: Cyclin-A and cyclin-E are regulators of G1-S phase of normal cell cycle. Integration of human papilloma virus high-risk (HR-HPV) could alter this mechanism, and its overexpression has been associated with poor prognosis in cervical cancer. AIM: To determine the expression of cyclin-A and cyclin-E, types of HR-HPV and physical state of DNA in cytologies with the diagnosis of low-grade squamous intraepithelial lesion (LSIL). MATERIALS AND METHODS: 115 cytological specimens in liquid base (liquid-PREP(™)) were analyzed. 25 specimens were with no signs of SIL (NSIL) and without HPV; 30 with NSIL with low-risk HPV (LR-HPV); 30 with NSIL with HR-HPV; and 30 with both LSIL and HR-HPV. The expression of cyclins was evaluated by immunocytochemistry; and the detection of viral DNA was done by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLPs) for genotyping or sequencing of HPV. The physical state of HPV was evaluated by in situ hybridization with amplification with tyramide. RESULTS: In the cytologies NSIL with LR-HPV, the expression of cyclin-A and cyclin-E was found respectively in 23.3% and 33.3% of the specimens. Among the specimens of NSIL with HR-HPV, 33.3% expressed cyclin-A and 40% cyclin-E, while 100% of the LSILs expressed the 2 cyclins. On the other hand, 100% of the samples NSIL with LR-HPV presented an episomal pattern. Of the specimens of NSIL with HR-HPV, 56.6% exhibited an episomal pattern, 23.3% integrated and 20%, mixed. Among the LSILs, 90% were mixed and 10% integrated. CONCLUSIONS: The cyclins A and E are present in the LSILs that occur predominantly in mixed state in the presence of HR-HPV.