Factors Defining Human Adipose Stem/Stromal Cell Immunomodulation in Vitro.

Human adipose tissue-derived stem/stromal cells (hASCs) are adult multipotent mesenchymal stem/stromal cells with immunomodulatory capacities. Here, we present up-to-date knowledge on the impact of different experimental and donor-related factors on hASC immunoregulatory functions in vitro. The experimental determinants include the immunological status of hASCs relative to target immune cells, contact vs. contactless interaction, and oxygen tension. Factors such as the ratio of hASCs to immune cells, the cellular context, the immune cell activation status, and coculture duration are also discussed. Conditioning of hASCs with different approaches before interaction with immune cells, hASC culture in xenogenic or xenofree culture medium, hASC culture in two-dimension vs. three-dimension with biomaterials, and the hASC passage number are among the experimental parameters that greatly may impact the hASC immunosuppressive potential in vitro, thus, they are also considered. Moreover, the influence of donor-related characteristics such as age, sex, and health status on hASC immunomodulation in vitro is reviewed. By analysis of the literature studies, most of the indicated determinants have been investigated in broad non-standardized ranges, so the results are not univocal. Clear conclusions cannot be drawn for the fine-tuned scenarios of many important factors to set a standard hASC immunopotency assay. Such variability needs to be carefully considered in further standardized research. Importantly, field experts' opinions may help to make it clearer.

Anti-Alzheimer Activity of Combinations of Cocoa with Vinpocetine or Other Nutraceuticals in Rat Model: Modulation of Wnt3/ß-Catenin/GSK-3ß/Nrf2/HO-1 and PERK/CHOP/Bcl-2 Pathways.

Alzheimer's disease (AD) is a devastating illness with limited therapeutic interventions. The aim of this study is to investigate the pathophysiological mechanisms underlying AD and explore the potential neuroprotective effects of cocoa, either alone or in combination with other nutraceuticals, in an animal model of aluminum-induced AD. Rats were divided into nine groups: control, aluminum chloride (AlCl3) alone, AlCl3 with cocoa alone, AlCl3 with vinpocetine (VIN), AlCl3 with epigallocatechin-3-gallate (EGCG), AlCl3 with coenzyme Q10 (CoQ10), AlCl3 with wheatgrass (WG), AlCl3 with vitamin (Vit) B complex, and AlCl3 with a combination of Vit C, Vit E, and selenium (Se). The animals were treated for five weeks, and we assessed behavioral, histopathological, and biochemical changes, focusing on oxidative stress, inflammation, Wnt/GSK-3ß/ß-catenin signaling, ER stress, autophagy, and apoptosis. AlCl3 administration induced oxidative stress, as evidenced by elevated levels of malondialdehyde (MDA) and downregulation of cellular antioxidants (Nrf2, HO-1, SOD, and TAC). AlCl3 also upregulated inflammatory biomarkers (TNF-α and IL-1ß) and GSK-3ß, leading to increased tau phosphorylation, decreased brain-derived neurotrophic factor (BDNF) expression, and downregulation of the Wnt/ß-catenin pathway. Furthermore, AlCl3 intensified C/EBP, p-PERK, GRP-78, and CHOP, indicating sustained ER stress, and decreased Beclin-1 and anti-apoptotic B-cell lymphoma 2 (Bcl-2) expressions. These alterations contributed to the observed behavioral and histological changes in the AlCl3-induced AD model. Administration of cocoa, either alone or in combination with other nutraceuticals, particularly VIN or EGCG, demonstrated remarkable amelioration of all assessed parameters. The combination of cocoa with nutraceuticals attenuated the AD-mediated deterioration by modulating interrelated pathophysiological pathways, including inflammation, antioxidant responses, GSK-3ß-Wnt/ß-catenin signaling, ER stress, and apoptosis. These findings provide insights into the intricate pathogenesis of AD and highlight the neuroprotective effects of nutraceuticals through multiple signaling pathways.

Potential role of PIM1 inhibition in the treatment of SARS-CoV-2 infection.

BACKGROUND: SARS-CoV-2 infection involves disturbing multiple molecular pathways related to immunity and cellular functions. PIM1 is a serine/threonine-protein kinase found to be involved in the pathogenesis of several viral infections. One PIM1 substrate, Myc, was reported to interact with TMPRSS2, which is crucial for SARS-CoV-2 cell entry. PIM1 inhibitors were reported to have antiviral activity through multiple mechanisms related to immunity and proliferation. This study aimed to evaluate the antiviral activity of 2-pyridone PIM1 inhibitor against SARS-CoV-2 and its potential role in hindering the progression of COVID-19. It also aimed to assess PIM1 inhibitor's effect on the expression of several genes of Notch signaling and Wnt pathways. In vitro study was conducted on Vero-E6 cells infected by SARS-CoV-2 "NRC-03-nhCoV" virus. Protein-protein interaction of the study genes was assessed to evaluate their relation to cell proliferation and immunity. The effect of 2-pyridone PIM1 inhibitor treatment on viral load and mRNA expression of target genes was assessed at three time points. RESULTS: Treatment with 2-pyridone PIM1 inhibitor showed potential antiviral activity against SARS-CoV-2 (IC50 of 37.255 µg/ml), significantly lowering the viral load. Functional enrichments of the studied genes include negative regulation of growth rate, several biological processes involved in cell proliferation, and Interleukin-4 production, with interleukin-6 as a predicted functional partner. These results suggest an interplay between study genes with relation to cell proliferation and immunity. Following in vitro SARS-CoV-2 infection, Notch pathway genes, CTNNB1, SUMO1, and TDG, were found to be overexpressed compared to uninfected cells. Treatment with 2-pyridone PIM1 inhibitor significantly lowers the expression levels of study genes, restoring Notch1 and BCL9 to the control level while decreasing Notch2 and CTNNB1 below control levels. CONCLUSION: 2-pyridone PIM1 inhibitor could hinder cellular entry of SARS-CoV-2 and modulate several pathways implicated in immunity, suggesting a potential benefit in the development of anti-SARS-CoV-2 therapeutic approach.

The role of PRDM1 gene polymorphism in the progression of hepatocellular carcinoma in Egyptian patients.

In Egypt, hepatocellular carcinoma (HCC) ranks as the second largest cause of cancer mortality. PRDM1 is a tumor suppressor gene essential for the differentiation and regulation activity of plasma cells and T cells. It plays a vital role in T cell exhaustion of chronic viral infection and HCC. We aimed to study the role of PRDM1 gene polymorphism in HCV and HCC-related to hepatitis C virus (HCV) progress in Egyptians. The case-control study included 300 Egyptian patients divided into 100 HCC,100 cirrhosis, and 100 control. Laboratory investigations were done for some clinicopathological biomarkers, including liver function tests, complete blood picture, serum alpha-fetoprotein, and hepatitis markers (HBsAg, anti-HCV-Ab). TaqMan allelic discrimination assay technique was used to genotype PRDM1 gene polymorphism. Multivariant analysis (logistic regression) assessed the association between the polymorphisms with HCC progression and designed the suggested model for HCC prediction. The frequencies of the G allele and GG phenotype in the control group were significantly more than that of the HCC and cirrhosis group. However, GA genotypes and A allele frequencies significantly increased in the HCC patients than in cirrhosis and controls. In addition, by comparing the HCC group and the non-HCC group (controls and cirrhotic patients), the subjects carrying AA or GA have 2 times more risk to develop HCC than those carrying GG genotypes (odd ratio = 2.045% and 95% confidence interval are (1.123-3.722) p = 0.019). Multivariate analysis results suggested a model of Aspartate transaminase (AST), Albumin, and PRDM1 polymorphism to predict the risk of HCC in Egyptians. In addition, PRDM1 polymorphism has an association with HCC prognosis (tumor size). For PRDM1 polymorphism, the A allele and AA might be considered as HCC-related to the HCV risk factor. In addition, AST, Albumin, and PRDM1 polymorphism predict the risk of HCC in Egyptians Therefore, the polymorphism might help in identifying the susceptible Egyptians to HCC. In addition, polymorphism might have a role in HCC prognosis.

MiR-155 and MiR-665 Role as Potential Non-invasive Biomarkers for Hepatocellular Carcinoma in Egyptian Patients with Chronic Hepatitis C Virus Infection.

BACKGROUND AND OBJECTIVES: Hepatocellular carcinoma (HCC) is the fourth leading cause of cancer associated death globally. Serum micro RNAs are full of potential as noninvasive biomarkers. Here, we aim to assess the performance of serum MicroRNA-155 and MicroRNA-665 as diagnostic biomarker for HCC comparing to AFP. METHODS: Serum samples were collected from 200 subjects (40 healthy control, 80 chronic hepatitis C patients with cirrhosis and without HCC (LC) and 80 HCC patients currently infected by hepatitis C infection and didn't start the treatment). The HCC patients didn't include alcoholic liver disease, nonalcoholic fatty liver disease nor autoimmune liver disease. MicroRNA-155 and MicroRNA-665 expression were measured by real-time quantitative PCR (RT-qPCR), while AFP level was assessed by ELISA method. RESULTS: Both miR-155 and miR-665 were significantly elevated in HCC group as compared to both control and LC groups. The comparison between LC and HCC patients revealed that the serum level of miR-155 was a significant increase in HCC patients compared to LC patients; however, the serum level of miR-665 didn't show any significant difference between the same two groups. MiR-665 expression level showed a direct correlation with tumor size in HCC patients. CONCLUSIONS: Using measurement against AFP level in serum, miR-665 is considered a promising serum biomarker for the diagnosis of HCC patients among the LC patients without HCC. MiR-155 didn't provide a better performance than serum AFP as a diagnostic biomarker among the same group. MiR-665 may serve as a good indicator for HCC prognosis.

Prognostic Significance of Serum Vitamin D Levels in Egyptian Females with Breast Cancer

Objective: To determine the frequency and prognostic significance of vitamin D deficiency in Egyptian women with breast cancer (BC). Methods: This prospective study included 50 women with primary invasive, non-metastatic BC. The serum level of 25-hydroxy vitamin D [25(OH)D was measured by ELISA at diagnosis, before any cancer treatment. Vitamin D deficiency was defined as 25(OH) D<20 ng/mL. Patients were followed up for a median of 30 months (range: 18-48). Results: The median level of 25(OH)D was 29.0 ng/mL (range: 10.0-55.0 ng/mL). Fifteen patients (30%) had vitamin D deficiency, which was positively associated with larger tumor size (p < 0.001), higher grade (p = 0.014), advanced stage (p = 0.001), lymph node positivity (p = 0.012), and HER2/neureceptor expression (p = 0.002). It was also linked with worse overall survival (OS) and disease free survival (DFS) (p = 0.026, and p = 0.004, respectively). On multivariate analysis, DFS was independently affected by vitamin D deficiency with an HR of 2.8 (95% CI: 1.6-7.0, p = 0.022) and advanced stage, i.e. stage II had worse survival compared to stage I with an HR of 4.8 (95%CI: 1.1-21.7, p = 0.042). Conclusion: Vitamin D deficiency had a negative effect on overall and disease-free survival in our breast cancer cases, being related to tumor size, stage, grade, nodal status and HER2/neu receptor expression.

Effect of Sirtuin 1 inhibition on matrix metalloproteinase 2 and Forkhead box O3a expression in breast cancer cells.

Breast cancer is the most common invasive cancer in women worldwide. Sirtuin 1 (SIRT1) has recently been shown to have implications in regulating cancer cell growth and apoptosis. SIRT1 regulates Forkhead box O3a (FOXO3a) by both inhibiting FOXO3-induced apoptosis and potentiating the ability of FOXO3a to resist oxidative stress. Matrix metalloproteinase 2 (MMP2) participates in tumor invasion and metastasis by degrading extracellular matrix. SIRT1 up regulates MMP2 expression by its deacetylation activity. This study aimed to investigate the expression of SIRT1, FOXO3a and MMP2 in breast tissues of women with breast cancer. In addition, the effect of SIRT1 inhibition on both FOXO3a and MMP2 expression in breast cancer (MCF-7) cells was assessed. The expression levels of SIRT1, FOXO3a and MMP2 in the breast tissues were determined by real-time PCR in 60 patients with malignant tumor and in 24 patients with benign tumors. After SIRT1 inhibition, protein levels of SIRT1 and FOXO3a were assessed by Western Blot and levels of MMP2 by ELISA in MCF-7 cells. The expression levels of SIRT1, FOXO3a and MMP2 were significantly higher in breast cancer tissues compared to in benign breast tumor and adjacent normal tissues. SIRT1, MMP2 and FOXO3a expression were associated directly with each other. SIRT1 inhibition suppresses MMP2 and FOXO3a expression compared to control MCF7. Sirtinol (SIRT1 inhibitor) effectively induced inhibition of MMP2 and FOXO3a expression in MCF-7 cells, indicating the promising therapeutic strategy of targeting SIRT1 for breast cancer.

Assessment of the -174G/C (rs1800795) and -572G/C (rs1800796) Interleukin 6 Gene Polymorphisms in Egyptian Patients with Rheumatoid Arthritis.

AIM: This study aimed to investigate genotype and allele frequencies of -174 (rs1800795) and -572 (rs1800796) IL-6 promoter gene polymorphisms in Egyptian patients with rheumatoid arthritis (RA) in comparison to control group. METHODS: The study was conducted on 198 Egyptian subjects (99 RA patients and 99 healthy control). The promoter region of the IL-6 gene was amplified by PCR using DNAs from patients and the controls, and their PCR products were digested by suitable enzymes. RESULTS: No statistical differences were found in -572G/C genotype (P = 0.177) or allele (P = 0.147) frequencies between RA patients and controls. Significant differences were observed in -174G/C genotype (P < 0.001) and allele (P < 0.001) frequencies between RA patients and controls. CONCLUSION: A significant association of IL-6 -174G/C gene polymorphism and RA in Egyptian population was found with significantly higher frequencies of GC and CC genotypes and C allele in RA patients compared to controls. No association was found between IL-6 -572G/C gene polymorphism and RA.

Ischemic heart diseases in Egypt: role of xanthine oxidase system and ischemia-modified albumin.

It is known that xanthine oxidoreductase contributes significantly to ischemia/reperfusion injury by generating reactive oxygen species. Ischemia-modified albumin (IMA) is a biomarker of acute myocardial ischemia with high sensitivity but moderate specificity. Our study aims to evaluate the xanthine oxidase (XO) system and the IMA level in the serum of patients with ischemic heart disease, and their correlation with traditional cardiac markers. The study was conducted on 60 patients with ischemic heart disease and 22 healthy subjects (control group). Subjects were divided into three groups: group I (30 patients with ST-elevated myocardial infarction), group II (30 patients with chronic stable angina), and the control group (22 subjects). The patients and controls had laboratory tests performed including lipid profile, cardiac enzymes, XO, uric acid, and IMA. The serum levels of XO and IMA were significantly higher in group I (1.65 ± 0.29 U/ml and 0.58 ± 0.15 ABSU, respectively) than in group II (1.11 ± 0.20 U/ml and 0.29 ± 0.10 ABSU, respectively) and the control group (0.95 ± 0.16 U/ml and 0.24 ± 0.08 ABSU, respectively) (P < 0.001). There was a significant positive correlation between XO and IMA in group I. Also, there was significant positive correlation between XO or IMA and other cardiac markers, with the highest level of significance between IMA and creatine kinase (CK-MB). In group II only XO activity was significantly elevated in comparison with controls. These results confirm the role of XO enzyme in ischemic heart disease with involvement of IMA, at a detectable level, during the early necrotic phase.