Favipiravir Protects Enterocytes From Cell Death After Inflammatory Storm.

Over the past years, inflammatory bowel disease (IBD) treatment has become more targeted, anticipating the use of immune-modifying therapies at an earlier stage. During the treatment process prevention and management of viral infections hold significant importance. The protective role of favipiravir on enterocytes which are affected by inflammation is still unknown. We aim to analyze the effects of favipiravir on enterocytes after an inflammatory condition. We conducted a 2,5-diphenyl-2H-tetrazolium bromide (MTT) assay to assess the cytotoxicity of favipiravir on intestinal epithelioid cells (IEC-6). To mimic the inflammation model in cell culture conditions, we exposed IEC-6 cells to tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ). The cells were categorized into four groups: control, inflammation model, application of favipiravir before inflammation (prophylactic), and application of favipiravir after inflammation (treatment). We assessed the presence and distribution of caspase 1, caspase 3, interleukin 6 (IL6), interleukin 8 (IL8), mixed lineage kinase domain-like protein (MLKL), receptor-interacting protein kinase 1 (RIPK1), and TNF-α using indirect immunoperoxidase staining. TNF-α and IL8 levels were analyzed with enzyme-linked immunosorbent assay (ELISA) in a culture medium. Caspase 1 was observed to be strong (+++) in the treatment group and weak (+) in the prophylactic group compared to the inflammation group. Caspase 3 was weak (+) in the inflammation group, and it was strong (+++) in the prophylactic and treatment group, the increase in the treatment group was significant. Therefore administering favipiravir before inducing inflammation appears to control the inflammatory caspase pathway in intestinal enterocytes, protecting them from inflammatory responses, while the caspase 3-dependent apoptotic pathway may not be active in enterocytes during inflammation. IL6 and IL8 were negative (-) in control, IL6 was weak (+) in inflammation and favipiravir treated groups; IL8 increased significantly in favipiravir groups compared to control and inflammation groups. Consequently, favipiravir may trigger IL6 release, initiating the inflammatory pathway and potentially enhancing IL8 interactions with other cytokines. TNF-α immunoreactivity was strong (+++) in the inflammation group, while it was moderate (++) in favipiravir-administered groups. MLKL immunoreactivity was strong (+++) in all groups, RIPK1 was weak (+) in control, strong (+++) in the inflammation and treatment group, moderate (++) in the prophylactic group, and the increase in inflammation and treatment group was significant compared to control. Our findings suggest that in the treatment group, necroptosis was triggered by increased MLKL and RIPK1, key players in inflammation and cell death. After immunocytochemical evaluation, our findings suggest that, after the onset of inflammation, favipiravir may play a role in cell death by increasing necroptosis rather than apoptosis.

The Role and Mechanisms of Action of Catechins in Neurodegenerative Diseases.

The prevalence, incidence and mortality rates of neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease are gradually increasing. New approaches are being developed to manage the progression and treatment of neurodegenerative diseases. Catechins, polyphenolic compounds, are key compounds that demonstrate therapeutic effects with their properties such as antioxidant, anti-inflammatory, anti-apoptotic properties in the prevention and treatment of neurodegenerative diseases. The therapeutic effects of catechins have been exhaustively studied in human and animal models. Catechins can have anti-inflammatory effects by suppressing inflammatory pathways and cytokines, as well as antioxidant effects such as chelating metal ions and scavenging radicals. They might reduce phosphorylation of tau proteins, aggregation of amyloid-beta and apoptotic proteins release. They can also decrease alpha-synuclein accumulation and increase dopamine levels. With all these effects, they can have an effect on neurodegenerative diseases. This review points to the potential mechanisms of catechins in neurodegenerative diseases, based on their findings in the literature review.Key teaching pointsCatechins can reduce amyloid-ß plaque aggregation and tau phosphorylation.Catechins can decrease alfa-synuclein levels.Catechins can protect neuronal cells with their anti-apoptotic effect.More comprehensive studies are needed to clarify this issue.

The role of resveratrol in diabetes and obesity associated with insulin resistance.

Diabetes mellitus is a significant health problem that is caused by chronic hyperglycaemia as a result of inadequate insulin production or ineffective insulin action in the body. In recent years, many new pharmacological and non-pharmacological therapies have been developed for improving pancreatic insulin secretion and insulin resistance. Resveratrol is a natural and biologically active stilbenoid polyphenol present in various plant species and has the potential to benefit diabetes. The anti-diabetic actions of resveratrol have also been extensively studied in diabetic human and animal models. Moreover, resveratrol might affect insulin sensitivity by regulating visceral fat derivated adipokine levels. The use of resveratrol in combination with anti-diabetic therapies or alone may have significant potential for the management of diabetes mellitus. This review provides an overview of the anti-diabetic action of resveratrol as well as the possible mechanisms that have an effect on insulin secretion and insulin resistance in diabetics.

Senescence-mediated anticancer effects of quercetin.

Cellular senescence plays a key role in aging and age-related disease initiation. It is a highly dynamic and multistep process that can be stimulated by various stimuli, including cellular stress, DNA damage, telomere shortening, and oncogene activation. Also, senescence is a potent antitumor mechanism, by preventing the proliferation of cancerous cells. However, some of the senescent cells have apoptosis resistance and can cause recurrence in cancer. A new class of drugs termed senolytics selectively kill and eliminate senescent cells. In recent years, natural compounds such as quercetin have been discovered to be effective as senolytic agents. Quercetin is a phytochemical that has strong antioxidant properties and pro-apoptotic effects and has been investigated for many years. Additionally, it has great potential to be used as a senolytic agent. According to preclinical and early-phase clinical data of senolytic agent research, quercetin administration appears to be effective in preventing or alleviating cancer formation. In this paper, we review the importance of cellular senescence in carcinogenesis and the effects of quercetin on senescence, as well as quercetin's potential effects as a pro-apoptotic agent and suppressor of cancer cell proliferation.

Neuroprotective effects of catechins in an experimental Parkinson's disease model and SK-N-AS cells: evaluation of cell viability, anti-inflammatory and anti-apoptotic effects.

OBJECTIVES: The aim of the study was to establish an in vitro Parkinson's disease (PD) model and to investigate the cell viability, anti-inflammatory, anti-apoptotic and neuroprotective effects of catechin and EGCG in SK-N-AS and in vitro PD model cells. METHOD: SK-N-AS human neuroblastoma cells were used. To develop an in vitro PD model, SK-N-AS cells were exposed to 6-hydroxydopamine. Model control was performed after ELISA analysis of dopamine and α-synuclein levels in the culture medium. Catechin and EGCG were administered to SK-N-AS and in vitro PD model cells. Cell viability was measured using MTT assay and trypan blue staining. Anti-inflammatory and anti-apoptotic activities of catechin and EGCG were investigated by indirect immunocytochemistry using anti-TNF-α, anti-IL-1ß and anti-caspase-3. RESULTS: After 24 hours of 6-hydroxydopamine administration at 50 µM, higher αlfa-synuclein and lower dopamine levels were found in PD model than SK-N-AS cells. Cell viability was similar between SK-N-AS and in vitro PD model cells. Treatment with both bioactive components increased cell viability of in vitro PD model cells. Caspase-3 immunoreactivity was significantly reduced in SK-N-AS and PD model cells after EGCG administration, while it was decreased only in PD model cells after catechin administration. IL-1ß staining intensity weakened after catechin administration in PD model cells, after EGCG administration in SK-N-AS cells. TNF-α staining intensity was similar in both cells. CONCLUSION: Catechin and EGCG increased cell viability in PD model neuron cells. Both components showed anti-apoptotic and anti-inflammatory effects. Catechin may be more effective in preventing damage to neurons PD.

Effect of kisspeptin-54 on ovarian levels of pigment epithelium-derived factor (PEDF) and vascular endothelial growth factor (VEGF) in an experimental model of ovarian hyperstimulation syndrome (OHSS).

The purpose of this study was to investigate the effect of kisspeptin-54 on ovarian morphology and vascular endothelial growth factor (VEGF), pigment epithelium-derived factor (PEDF), protein kinase A (PKA) and protein kinase C (PKC) levels in an ovarian hyperstimulation syndrome (OHSS) rat model, which is a possible complication of controlled ovarian hyperstimulation. For this purpose, immature female Sprague-Dawley rats (25days old, 30-40g) were randomly divided into five groups (control, sham, OHSS model, short-term kisspeptin-54 administered OHSS model and long-term kisspeptin-54 administered OHSS model). Serum LH and FSH levels were determined by enzyme-linked immunosorbent assay. Immunohistochemistry and quantitative RT-PCR were performed for VEGF, PEDF, PKA and PKC in ovaries and granulosa cells, respectively. It was observed that there was dilatation in fallopian tubes and an abnormal increase in ovarian weight and volume in the OHSS group, and these morphologies decreased with kisspeptin-54 treatment. After the administration of kisspeptin-54 in the OHSS group, VEGF, PKA and PKC levels reduced and PEDF levels increased in both mRNA and protein levels. It was determined that in the OHSS model, VEGF increased as PEDF decreased, and kisspeptin-54 reduced the effects of OHSS. It was determined that long-term kisspeptin-54 treatment was more effective than short-term administration. It is considered that kisspeptin-54 is an agent that protects ovarian reserve and oocyte maturation in women at risk of OHSS.

Synthesis and Apoptotic Activities of New 2(3H)-benzoxazolone Derivatives in Breast Cancer Cells.

BACKGROUND: 2(3H)-Benzoxazolone derivatives are preferential structural blocks in pharmacological probe designing with the possibility of modifications at various positions on the core structure. Benzoxazolones showed various biological activities such as analgesics, anti-inflammatory and anti-cancer. OBJECTIVE: In the present work, we have prepared new Mannich bases of 2(3H)-benzoxazolone derivatives and evaluated their cytotoxicities and proapoptotic properties in MCF-7 breast cancer cell line. METHODS: The structures of these compounds were characterized by FT-IR, elemental analysis, 1H and 13C NMR. Cytotoxicities of all the target compounds were investigated by MTT assay. Apoptotic properties of compounds were evaluated by immunocytochemistry using antibodies against caspase-3, cytochrome-c, FasL, and also TUNEL assay. RESULTS: These two novel compounds, 1 and 2, both have the same piperazine substituent on the nitrogen atom of benzoxazolone and the main difference in the structures of these compounds is the presence of Cl substituent at the 5- position of the benzoxazolone ring. MTT results showed that compounds 1 and 2 were effective in terms of reduction of cell viability at 100µM and 50µM concentration for 48h, respectively. As a result of immunohistochemical staining, Fas L and caspase-3 immunoreactivities were significantly increased in MCF-7 cells after treatment with compound 1. Additionally, caspase-3 and cytochrome-c immunoreactivities were also increased significantly in MCF-7 cells after treatment with compound 2. The number of TUNEL positive cells was significantly higher in MCF-7 cells when compared with the control group after treatment with both compounds 1 and 2. CONCLUSION: It could be concluded that N-substituted benzoxazolone derivatives increase potential anti-cancer effects and they could be promising novel therapeutic agents for chemotherapy.

The correlation between unexplained infertility and exosomes.

OBJECTIVES: Endometrial receptivity plays the most important role for successful implantation. Increasing endometrial receptivity may improve infertility and increase Assisted Reproductive Technologies success. The aim of this study was to investigate the effect of exosome specific markers CD63 and CD9 which are promising molecules in the pathogenesis and treatment of many diseases on endometrial receptivity in women with unexplained infertility. MATERIAL AND METHODS: This prospective study was conducted between November 2015 and March 2017. Proliferation and secretion periods of endometrial samples from fertile and infertile cases were collected. The paraffin-embedded tissue sections were stained with hematoxylin-eosin for the immunohistochemical analysis distributions of CD63 and CD9. RESULTS: The results of this study demonstrated that the CD63 immunoreactivity was higher in both luminal and glandular epithelium of infertile patients when compared with fertile patients during the proliferative phase (p = 0.009, p = 0.008). In the infertile proliferation phase, endometrium CD9 immunoreactivity was rarely detected in both the luminal and glandular epithelium. In the secretion phase of endometrium, CD9 immunoreactivity was mild in fertile patients, the increased immunoreactivity of CD9 was observed in both luminal and glandular epithelium of infertile patients (p = 0.037, p = 0.037). CONCLUSIONS: Increased levels of CD63 in infertile proliferation phase endometrium should represent an unfavorable signaling. Moreover, the increased levels of CD9 in infertile secretion phase endometrium could be used as a biomarker to evaluate endometrial receptivity. These exosome-specific markers can be considered as potential molecular markers of infertility.

The Effect of Resveratrol and Quercetin on Epithelial-Mesenchymal Transition in Pancreatic Cancer Stem Cell.

Resveratrol and quercetin are phytochemicals that are found in a variety of plants. The aim of this study was to investigate the effect of resveratrol and quercetin on epithelial-mesenchymal transition (EMT) of CD133+ and CD133- pancreatic cancer cells. Cancer stem cells (CD133+ cells) were obtained from the PANC-1 cells by the MiniMACS system. CD133+ and CD133- PANC-1 cells were treated with different concentrations (5, 10, 25, 50, and 100 µM) of resveratrol and quercetin. Cell growth and cytotoxicity were evaluated by MTT assay. Anticancer and anti-metastatic properties of resveratrol and quercetin were determined by immunocytochemistry using antibodies (ACTA-2, IL-1ß, N-cadherin, TNF-α, and vimentin). The immunostaining intensity of CD133+ cells was stronger than CD133- cells. ACTA-2, IL-1ß, and N-cadherin immunoreactivities were significantly decreased, whereas TNF-α and vimentin immunoreactivities significantly increased in quercetin-treated CD133+ cells. Moreover, N-cadherin and TNF-α immunoreactivities significantly decreased in resveratrol-treated CD133+ cells. The reduction in N-cadherin and ACTA-2 immunoreactivities was higher than the increase in vimentin immunoreactivity, quercetin could prevent EMT to a greater extent than resveratrol in pancreatic cancer stem cells because of the reduced expression of N-cadherin. Quercetin could be more effective in inhibiting metastasis compared to resveratrol.

Effects of 5-fluorouracil and gemcitabine on a breast cancer cell line (MCF-7) via the JAK/STAT pathway.

Aberrant activation of the JAK/STAT pathway may predispose to malignancy as a consequence of the deregulation of cell proliferation, differentiation or apoptosis such as in cancer of the blood, head and neck, and breast. In our study we aimed to investigate the effects of 5-fluorouracil (5-FU) and gemcitabine on a breast cancer cell line (MCF-7 cells) via the JAK/STAT pathway. Distribution of JAK1, JAK2, JAK3 and STAT2, STAT3, STAT4, STAT5 were evaluated on MCF-7 cells following gemcitabine and 5-FU treatment and in the absence of drug treatment by an indirect immunohistochemical method. It was observed that JAK1, JAK3, STAT5 and particularly STAT2 activation were more effective than the other JAK/STATs in breast cancer progression. Following treatment with 5-FU, JAK1 and STAT5 immunoreactivities were decreased in MCF-7 cells in comparison with both gemcitabine-treated and non-treated groups. These results suggest that the JAK/STAT pathway plays an important role in breast cancer pathogenesis and may be more affected after 5-FU treatment rather than gemcitabine. Drugs which block STAT5 may provide a novel therapeutic approach for the treatment of breast cancer.

The effects of Gemcitabine and Vinorelbine on inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS) distribution of MCF-7 breast cancer cells.

Gemcitabine, which induces S-phase arrest, and Vinorelbine, which arrests microtubule organization, are two agents that have demonstrate preferred anti-tumor activity. Nitric oxide acts in diverse functions including anti-tumor and anti-pathogenic activities. In this study, we aimed to examine the distribution of immunoreactivities of inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS) in cells of the MCF-7 breast cancer cell line in response to treatment with Gemcitabine (G), Vinorelbine (V) and combination of Gemcitabine and Vinorelbine (G+V). The distributions of iNOS and eNOS were determined by using indirect immunoperoxidase or immunofluorescence methods and ELISA. Cells incubated with G, V and G+V for 24, 48 and 72h were immunolabelled with anti-eNOS and anti-iNOS primary antibodies. Apoptosis was determined by TUNEL assay. A significant increase of eNOS immunolabelling on MCF-7 cells treated with G and G+V was observed. Apoptotic cells were also detected in G, V and G+V treated MCF-7 cells. The immunolabelling of iNOS was detected in all groups but this immunoreactivity was not different among the groups. In conclusion, while G treatment, induced S-phase arrest, triggered the NOS pathway after treatment of MCF-7 cells, V treatment, arrested microtubule organization and did not change the NOS pathway. Detection of increased eNOS immunolabelling and apoptosis after G treatment of MCF-7 cells could be important to the treatment of human breast cancer.

The maturity of intestinal neomucosa: integrin expression and ultrastructural aspects.

BACKGROUND/PURPOSE: The maturity of neomucosa growing on a serosal surface for the treatment of short bowel syndrome still is questionable. The aim of this study was to evaluate the intestinal neomucosa to assess its histologic maturity. METHODS: A 6-cm-long isolated ileal segment (IS) was prepared in 8 Wistar albino-type rats. The IS was divided from the antimesenteric side, and 2 intestinal tubes were established, which shared a common wall and a common pedicle. After ileal biopsy sampling for the control group (CG), the IS was fashioned into a mucous fistula. Eight weeks later, all the rats were killed, and the ISs were investigated for neomucosal growth. Sections were prepared with periodic acid shift (PAS) and H & E staining for light microscopy. They also were evaluated by transmission electron microscopy. The microscopic morphology of the 2 groups was evaluated. Immunohistochemical staining was performed to show the expression of the tissue beta1, alpha3 and alpha2beta1 integrin subunits of both the neomucosa (NS) and control group (CG) segments. RESULTS: Sections of the NS showed a well-arranged columnar epithelial cell layer with goblet cells that were generally located superficially and with a complete basement membrane. Under the electron microscope, the sections from the NS group showed an epithelial cell layer with proper microvilli of the same height, although they were shorter than those of the CG, and tight intercellular junctions between the epithelial cells. Significant differences between the NS and CG groups were found in the measurements of villus width at base, microvillus surface, and microvillus height. The lamina propria consisted of rich collagen fibers and active fibroblasts in the NS group. In the immunohistochemical staining, although beta1 integrine showed a dense distribution (+++) in the lamina propria, particularly localizing at the depth of the tunica mucosa layer, alpha3 integrin was observed to have a less dense immunoreactivity (++) in both groups. The expression of alpha2beta1 integrin showed slight and dispersed (+) staining. CONCLUSIONS: The NS showed histologic maturity and ultimate structural similarity with the native small bowel mucosa, which provides strong indirect evidence for the proper functioning of the neomucosa.