HLA alleles, haplotypes frequencies, and their association with hematological disorders: a report from 1550 families whose patients underwent allogeneic bone marrow transplantation in Egypt.

HLA alleles are representative of ethnicities and may play important roles in predisposition to hematological disorders. We analyzed DNA samples for HLA-A, -B, -C, -DRB1, and -DQB1 loci, from 1550 patients and 4450 potential related donors by PCR-SSO (Polymerase chain reaction sequence-specific oligonucleotides) and estimated allele frequencies in donors and patients from 1550 families who underwent bone marrow transplantation (BMT) in Egypt. We also studied the association between HLA allele frequencies and incidence of acute myeloid leukemia, acute lymphoblastic leukemia, and severe aplastic anemia. The most frequently observed HLA class I alleles were HLA- A*01:01 (16.9%), A*02:01 (16.1%), B*41:01 (8.7%), B*49:01 (7.3%), C*06:02 (25.1%), and C*07:01 (25.1%), and the most frequently observed class II alleles were HLA-DRB1*11:01 (11.8%), DRB1*03:01 (11.6%), DQB1*03:01 (27.5%), and DQB1*05:01 (18.9%). The most frequently observed haplotypes were A*33:01~B*14:02 ~ DRB1*01:02 (2.35%) and A*01:01~B*52:01~DRB1*15:01 (2.11%). HLA-DRB1*07:01 was associated with higher AML odds (OR, 1.26; 95% CI, 1.02-1.55; p = 0.030). Only HLA-B38 antigen showed a trend towards increased odds of ALL (OR, 1.52; 95% CI, 1.00-2.30; p = 0.049) HLA-A*02:01, -B*14:02, and -DRB1*15:01 were associated with higher odds of SAA (A*02:01: OR, 1.35; 95% CI, 1.07-1.70; p = 0.010; B*14:02: OR, 1.43; 95% CI, 1.06-1.93; p = 0.020; DRB1*15:01: OR, 1.32; 95% CI, 1.07-1.64; p = 0.011). This study provides estimates of HLA allele and haplotype frequencies and their association with hematological disorders in an Egyptian population.

FBLN2 is associated with basal cell markers Krt14 and ITGB1 in mouse mammary epithelial cells and has a preferential expression in molecular subtypes of human breast cancer.

BACKGROUND: Fibulin-2 (FBLN2) is a secreted extracellular matrix (ECM) glycoprotein and has been identified in the mouse mammary gland, in cap cells of terminal end buds (TEBs) during puberty, and around myoepithelial cells during early pregnancy. It is required for basement membrane (BM) integrity in mammary epithelium, and its loss has been associated with human breast cancer invasion. Herein, we attempted to confirm the relevance of FBLN2 to myoepithelial phenotype in mammary epithelium and to assess its expression in molecular subtypes of human breast cancer. METHODS: The relationship between FBLN2 expression and epithelial markers was investigated in pubertal mouse mammary glands and the EpH4 mouse mammary epithelial cell line using immunohistochemistry, immunocytochemistry, and immunoblotting. Human breast cancer mRNA data from the METABRIC and TCGA datasets from Bioportal were analyzed to assess the association of Fbln2 expression with epithelial markers, and with molecular subtypes. Survival curves were generated using data from the METABRIC dataset and the KM databases. RESULTS: FBLN2 knockdown in mouse mammary epithelial cells was associated with a reduction in KRT14 and an increase in KRT18. Further, TGFß3 treatment resulted in the upregulation of FBLN2 in vitro. Meta-analyses of human breast cancer datasets from Bioportal showed a higher expression of Fbln2 mRNA in claudin-low, LumA, and normal-like breast cancers compared to LumB, Her2 +, and Basal-like subgroups. Fbln2 mRNA levels were positively associated with mesenchymal markers, myoepithelial markers, and markers of epithelial-mesenchymal transition. Higher expression of Fbln2 mRNA was associated with better prognosis in less advanced breast cancer and this pattern was reversed in more advanced lesions. CONCLUSION: With further validation, these observations may offer a molecular prognostic tool for human breast cancer for more personalized therapeutic approaches.

Detection of MET gene somatic mutations in hepatocellular carcinoma of Egyptian patients using next-generation sequencing.

INTRODUCTION: Hepatocellular carcinoma (HCC) is the sixth most common type of cancer worldwide and fourth in Egypt. Liquid biopsy is important to get cell-tumour DNA (ctDNA), for subsequent utilisation as a biomarker for cancer diagnosis, prognosis, and treatment. In clinical oncology, ctDNA analysis is utilised in cancer screening. METHODS: The collected 48 blood samples from HCC patients were classified according to Barcelona Clinic Liver Cancer (BCLC) staging, in addition to Hepatitis C Virus (HCV) group and normal group. After the liquid biopsy, ctDNA and genomic DNA (gDNA) of the same individual were extracted. Next-generation sequencing (NGS) was conducted using a Hot spot panel, and data analysis via different cancer databases was performed. RESULTS: There were no significant differences in the detected mutation frequency between groups. The frequency of mutations was higher in ctDNA than in the gDNA samples from the same patients. Hence, it can be concluded that these mutations are somatic mutations, rather than germline mutations. CONCLUSION: Screening of the targeted genes such as c-MET for potential mutations is very important in the determination of the appropriate therapy. Therefore, it can be used as a biomarker in the prognosis of HCC. Such screenings are also of paramount importance in the development of personalised medicine.

The Transgene Expression of the Immature Form of the HCV Core Protein (C191) and the LncRNA MEG3 Increases Apoptosis in HepG2 Cells.

Long non-coding RNAs (lncRNAs) are regulated in cancer cells, including lncRNA MEG3, which is downregulated in Hepatocellular Carcinoma (HCC). In addition, hepatitis C virus (HCV) core proteins are known to dysregulate important cellular pathways that are linked to HCC development. In this study, we were interested in evaluating the overexpression of lncRNA MEG3, either alone or in combination with two forms of HCV core protein (C173 and C191) in HepG2 cells. Cell viability was assessed by MTT assay. Transcripts' levels of key genes known to be regulated in HCC, such as p53, DNMT1, miRNA152, TGF-b, and BCL-2, were measured by qRT-PCR. Protein expression levels of caspase-3 and MKI67 were determined by immunocytochemistry and apoptosis assays. The co-expression of lncRNA MEG3 and C191 resulted in a marked increase and accumulation of dead cells and a reduction in cell viability. In addition, a marked increase in the expression of tumor suppressor genes (p53 and miRNA152), as well as a marked decrease in the expression of oncogenes (DNMT1, BCL2, and TGF-b), were detected. Moreover, apoptosis assay results revealed a significant increase in total apoptosis (early and late). Finally, immunocytochemistry results detected a significant increase in apoptotic marker caspase-3 and a decrease in tumor marker MKI67. In this study, transgene expression of C191 and lncRNA MEG3 showed induction in apoptosis in HepG2 cells greater than the expression of each one alone. These results suggest potential anticancer characteristics.

The regulatory role of long non- coding RNAs as a novel controller of immune response against cancer cells.

Immunotherapy has been established as a promising therapy for different cancer types. However, many patients experience primary or secondary resistance to treatment. Immune cells and anti-inflammatory factors are regulated by long noncoding RNAs (lncRNAs). In addition, lncRNAs have a role in immune resistance through antigen presentation loss or attenuation, PD-L1 upregulation, loss of T-cell activities, and activation of G-MDSCs and Tregs in the tumor environment. LncRNAs can also influence the interaction between cancer stem cells and immune cells in the tumor microenvironment, potentially resulting in cancer stem cell resistance to immunotherapy. Immunological-related lncRNAs can influence immune responses either directly by affecting neighboring protein-coding genes or indirectly by sponging miRNAs through various mechanisms. We have emphasized the role and levels of expression of lncRNAs that have been linked to immune cell formation, differentiation, and activation, which may have an influence on immunotherapy efficacy.

Role of adipose tissue-derived cytokines in the progression of inflammatory breast cancer in patients with obesity.

BACKGROUND: Inflammatory breast cancer (IBC) represents a deadly aggressive phenotype of breast cancer (BC) with a unique clinicopathological presentation and low survival rate. In fact, obesity represents an important risk factor for BC. Although several studies have identified different cellular-derived and molecular factors involved in IBC progression, the role of adipocytes remains unclear. Cancer-associated adipose tissue (CAAT) expresses a variety of adipokines, which contribute to tumorigenesis and the regulation of cancer stem cell (CSC). This research investigated the potential effect of the secretome of CAAT explants from patients with BC on the progression and metastasis of the disease. METHODS: This study established an ex-vivo culture of CAAT excised from IBC (n = 13) vs. non-IBC (n = 31) patients with obesity and profiled their secretome using a cytokine antibody array. Furthermore, the quantitative PCR (qPCR) methodology was used to validate the levels of predominant cytokines at the transcript level after culture in a medium conditioned by CAAT. Moreover, the impact of the CAAT secretome on the expression of epithelial-mesenchymal transition (EMT) and cells with stem cell (CSC) markers was studied in the non-IBC MDA-MB-231 and the IBC SUM-149 cell lines. The statistical differences between variables were evaluated using the chi-squared test and unpaired a Student's t-test. RESULTS: The results of cytokine array profiling revealed an overall significantly higher level of a panel of 28 cytokines secreted by the CAAT ex-vivo culture from IBC patients with obesity compared to those with non-IBC. Of note, interleukin-6 (IL-6), interleukin-8 (IL-8), and monocyte chemo-attractant protein 1 (MCP-1) were the major adipokines secreted by the CAAT IBC patients with obesity. Moreover, the qPCR results indicated a significant upregulation of the IL-6, IL-8, and MCP-1 mRNAs in CAAT ex-vivo culture of patients with IBC vs. those with non-IBC. Intriguingly, a qPCR data analysis showed that the CAAT secretome secretions from patients with non-IBC downregulated the mRNA levels of the CD24 CSC marker and of the epithelial marker E-cadherin in the non-IBC cell line. By contrast, E-cadherin was upregulated in the SUM-149 cell. CONCLUSIONS: This study identified the overexpression of IL-6, IL-8, and MCP-1 as prognostic markers of CAAT from patients with IBC but not from those with non-IBC ; moreover, their upregulation might be associated with IBC aggressiveness via the regulation of CSC and EMT markers. This study proposed that targeting IL-6, IL-8, and MCP-1 may represent a therapeutic option that should be considered in the treatment of patients with IBC.

Association of vitamin D receptor gene polymorphisms with breast cancer risk in an Egyptian population.

This study aimed to explore whether genetic polymorphisms in vitamin D receptor are correlated to the breast cancer prevalence in an Egyptian population. Polymerase chain reaction-restriction fragment polymorphism was used to genotype three frequently analyzed vitamin D receptor gene single-nucleotide polymorphisms (rs1544410, rs7975232, and rs731236) and were identified by sequencing analysis. This is the first study that recorded a new single-nucleotide polymorphism in ApaI genotype within an Egyptian population and was registered with the accession number KY859868. The authors found that TC in rs731236, and TG in KY859868 single-nucleotide polymorphism showed significant distribution differences with an increased risk of breast cancer ( p < 0.05, odds ratio = 3.71, 95% confidence interval: 1.04-13.28 and p < 0.001, odds ratio = 7.05, 95% confidence interval: 2.02-24, respectively) compared with the wild-type TT genotype carriers in both single-nucleotide polymorphisms. In addition, the distribution frequencies of haplotypes ACT, GTT, and ATT in the patients group were significant, where ATT haplotype was associated with the highest breast cancer risk among all other haplotypes in the patients group ( p = 0.0023, odds ratio = 1.72, 95% confidence interval: 1.24-2.437). In conclusion, vitamin D receptors ApaI and TaqI confer high breast cancer susceptibility, particularly in Egyptians females carrying haplotype ATT. However, further studies focusing on the vitamin D receptor variants and haplotypes effects on vitamin D and vitamin D receptor concentrations, activities, and functionalities are needed.

Inflammatory breast cancer: High incidence of GCC haplotypes (-1082A/G, -819T/C, and -592A/C) in the interleukin-10 gene promoter correlates with over-expression of interleukin-10 in patients' carcinoma tissues.

Interleukin-10 is involved in carcinogenesis by supporting tumor escape from the immune response. The aim of this study was to assess the single nucleotide polymorphisms, -1082A/G, -819T/C and -592A/C, in interleukin-10 gene promoter in inflammatory breast cancer compared to non-inflammatory breast cancer and association of these polymorphisms with interleukin-10 gene expression. We enrolled 105 breast cancer tissue (72 non-inflammatory breast cancer and 33 inflammatory breast cancer) patients and we determined the three studied single nucleotide polymorphisms in all samples by polymerase chain reaction restriction fragment length polymorphism and investigated their association with the disease and with various prognostic factors. In addition, we assessed the expression of interleukin-10 gene by real-time quantitative reverse transcription polymerase chain reaction and the correlation between studied single nucleotide polymorphisms and interleukin-10 messenger RNA expression. We found co-dominant effect as the best inheritance model (in the three studied single nucleotide polymorphisms in non-inflammatory breast cancer and inflammatory breast cancer samples), and we didn't identify any association between single nucleotide polymorphisms genotypes and breast cancer prognostic factors. However, GCC haplotype was found highly associated with inflammatory breast cancer risk (p < 0.001, odds ratio = 43.05). Moreover, the expression of interleukin-10 messenger RNA was significantly higher (p < 0.001) by 5.28-fold and 8.95-fold than non-inflammatory breast cancer and healthy control, respectively, where GCC haplotype significantly increased interleukin-10 gene expression (r = 0.9, p < 0.001).

Chalcones: Promising therapeutic agents targeting key players and signaling pathways regulating the hallmarks of cancer.

The need for innovative anticancer treatments with high effectiveness and low toxicity is urgent due to the development of malignancies that are resistant to chemotherapeutic agents and the poor specificity of existing anticancer treatments. Chalcones are 1,3-diaryl-2-propen-1-ones, which are the precursors for flavonoids and isoflavonoids. Chalcones are readily available from a wide range of natural resources and consist of very basic chemical scaffolds. Because the ease with which the synthesis it allows for the production of several chalcone derivatives. Various in-vitro and in-vivo studies indicate that naturally occurring and synthetic chalcone derivatives exhibit promising biological activities against cancer hallmarks such as proliferation, angiogenesis, invasion, metastasis, inflammation, stemness, and regulation of cancer epigenetics. According to their structure and functional groups, chalcones derivatives and their hybrid compounds exert a broad range of biological activities through targeting key elements and signaling molecules relevant to cancer progression. This review will provide valuable insights into the latest updates of chalcone groups as anticancer agents and extensively discuss their underlying molecular mechanisms of action.

Tissue Distribution, Histopathological and Genotoxic Effects of Magnetite Nanoparticles on Ehrlich Solid Carcinoma.

Nanoparticles can potentially cause adverse effects on cellular and molecular level. The present study aimed to investigate the histopathological changes and DNA damage effects of magnetite nanoparticles (MNPs) on female albino mice model with Ehrlich solid carcinoma (ESC). Magnetite nanoparticles coated with L-ascorbic acid (size ~ 25.0 nm) were synthesized and characterized. Mice were treated with MNPs day by day, intraperitoneally (IP), intramuscularly (IM), or intratumorally (IT). Autopsy samples were taken from the solid tumor, thigh muscle, liver, kidney, lung, spleen, and brain for assessment of iron content, histopathological examination, and genotoxicity using comet assay. The liver, spleen, lung, and heart had significantly higher iron content in groups treated IP. On the other hand, tumor, muscles, and the liver had significantly higher iron content in groups treated IT. MNPs induced a significant DNA damage in IT treated ESC. While a significant DNA damage was detected in the liver of the IP treated group, but no significant DNA damage could be detected in the brain. Histopathological findings in ESC revealed a marked tumor necrosis, 50% in group injected IT but 40% in group injected IP and 20% only in untreated tumors. Other findings include inflammatory cell infiltration, dilatation, and congestion of blood vessels of different organs of treated groups in addition to appearance of metastatic cancer cells in the liver of non-treated tumor group. MNPs could have an antitumor effect but it is recommended to be injected intratumorally to be directed to the tumor tissues and reduce its adverse effects on healthy tissues.

[1,2,4] Triazolo [3,4-a]isoquinoline chalcone derivative exhibits anticancer activity via induction of oxidative stress, DNA damage, and apoptosis in Ehrlich solid carcinoma-bearing mice.

Despite the advances made in cancer therapeutics, their adverse effects remain a major concern, putting safer therapeutic options in high demand. Since chalcones, a group of flavonoids and isoflavonoids, act as promising anticancer agents, we aimed to evaluate the in vivo anticancer activity of a synthetic isoquinoline chalcone (CHE) in a mice model with Ehrlich solid carcinoma. Our in vivo pilot experiments revealed that the maximum tolerated body weight-adjusted CHE dose was 428 mg/kg. Female BALB/c mice were inoculated with Ehrlich ascites carcinoma cells and randomly assigned to three different CHE doses administered intraperitoneally (IP; 107, 214, and 321 mg/kg) twice a week for two consecutive weeks. A group injected with doxorubicin (DOX; 4 mg/kg IP) was used as a positive control. We found that in CHE-treated groups: (1) tumor weight was significantly decreased; (2) the total antioxidant concentration was substantially depleted in tumor tissues, resulting in elevated oxidative stress and DNA damage evidenced through DNA fragmentation and comet assays; (3) pro-apoptotic genes p53 and Bax, assessed via qPCR, were significantly upregulated. Interestingly, CHE treatment reduced immunohistochemical staining of the proliferative marker ki67, whereas BAX was increased. Notably, histopathological examination indicated that unlike DOX, CHE treatment had minimal toxicity on the liver and kidney. In conclusion, CHE exerts antitumor activity via induction of oxidative stress and DNA damage that lead to apoptosis, making CHE a promising candidate for solid tumor therapy.

Silica-coated graphene compared to Si-CdSe/ZnS quantum dots: Toxicity, emission stability, and role of silica in the uptake process for imaging purposes.

Quantum dots (QDs) present a special type of nanocrystals (NCs) due to their unique optical and chemical properties. While cadmium-based QDs (Cd-QDs) have the most favorable physicochemical properties, their toxicity, instability in the aqueous phase, and loss of brightness at high temperature are some of the obstacles that prevent the wide use of Cd-QDs. Carbon-based QDs as graphene quantum dots (GQDs) represent a very promising biocompatible replacement. In the present work, we mainly focus on comparing the efficiency and uptake of GQDs and Cd-QDs for fluorescent imaging purposes and studying the effect of growing silica shell on the emission and the uptake of QDs inside living human and bacterial cells. Graphene and CdSe/ZnS QDs were prepared and encapsulated in silica to increase their emission and uptake by living cells. Moreover, we studied their photostability and cytotoxicity. The Prepared G-Si QDs showed good emission inside the cytoplasmic portion of the liver hepatocellular carcinoma cell line (HepG2) and Bacillus subtilis (B. subtilis), but they revealed lower photoluminescence (PL) intensity compared to Si-CdSe/ZnS NCs although G-Si QDs are advantageous in other aspects, i.e. possess lower toxicity and higher stability with temperature variations.

Combining Bevacizumab with Knocked-Down ß-Catenin Reduces VEGF-A and Slug mRNA in HepG2 but not in Caco-2 Cell Lines.

BACKGROUND: Bevacizumab (Bev) resistance is hypothesized to be overcome by combining inhibitors of other signalling pathways. OBJECTIVE: We aimed to study the effect of combining Bev with knocked down ß-catenin (Bev-ß-cat-siRNA) on the expression of VEGF-A, Slug, NFκB, and its two target genes, c-Flip and FasR, in HepG2. Expression of VEGF-A and Slug was also studied in Caco-2 cells. METHODS: Cultured cells were divided into six groups 1) cells treated with Bev, 2) cells treated with ß-catenin-siRNA, 3) cells treated with Bev-ß-cat-siRNA, 4) cells treated with negative control, 5) cells treated with Bev-negative control, and 6) untreated cells. Expressions were assessed using qPCR and western blotting. RESULTS: Bev-ß-cat-siRNA significantly reduced the mRNA level of VEGF-A, which was initially increased in response to Bev alone in HepG2 but not in Caco-2. Additionally, Bev-ß-cat-siRNA significantly decreased Slug mRNA level compared to Bev treated HepG2 cells. In contrast, VEGF-A and Slug mRNA levels in Bev group were remarkably lower than Bev-ß-cat-siRNA in Caco-2 cells. Distinct ß-catenin and Slug protein expressions were noticed in HepG2 and Caco-2 cells. On the other hand, Bev-ß-catsiRNA remarkably reduced the level of NFκB, FasR, and c-Flip compared to Bev treated HepG2 cells, although the difference was not statistically significant. CONCLUSION: We conclude that combining Bevacizumab with knocked down ß-catenin reduces the expression of VEGF-A and Slug in HepG2 but not in Caco-2 cells.

High Frequency of ASXL1 and IDH Mutations in Young Acute Myeloid Leukemia Egyptian Patients.

BACKGROUND: Prognostication of AML patients depends on association of genetic and epigenetic abnormalities. We aimed to evaluate the frequency and prognostic significance of Additional Sex comb's Like1 (ASXL1), Isocitrate Dehydrogenase (IDH) and Casitas B- lineage Lymphoma (CBL) mutations in AML assessing their association with different cytogenetic risk category. METHODS: We used High Resolution Melting (HRM) technology that detects small differences in PCR amplified sequences by direct melting using EvaGreen saturating dye to analyze epigenetic mutations in 70 denovo AML patients. RESULTS: Median age of AML patients was 39.5 years (18-75). ASXL1, IDH and CBL mutations were detected in 14 (20%), 10 (14%) and 5 (7%) patients, respectively. Mean age of ASXL1 and IDH mutants vs. wild type was 35.9±14.6 years and 42.9±14.4 years (p=0.114) and 46.7±15.2 years vs. 40.6±14.5 years (p=0.290), respectively. AML cytogenetic risk groups included low (25/70, 36%), intermediate (33/70, 47%) and high-risk (12/70, 17%). Nine/14 (64%) ASXL1 and 8/10 (80%) IDH mutants were classified as intermediate risk and 9 ASXL1 positive (64%) were adolescent and young adults (AYA). Overall survival (OS) of mutant ASXL1 vs. wild type was 1.1 years (95% CI 0.83-1.4) vs. 1.9 years (95% CI 0.71-7.51), respectively (p=0.056). OS of mutant IDH vs. wild type was 1.25 years (95% CI 0.85-1.6) vs. 1.8 years (95% CI 1.2-6.7), respectively (p=0.020). In intermediate risk cytogenetic group, ASXL1 and IDH mutants had shorter OS than wild type; 1.1 years (95% CI 0.97-1.2) vs. 2.1 years (95% CI 0.14-10.8) (p=0.002) and 1.8 years (95% CI 0.69-3.15) vs. 2.3 years (95% CI 1.1-5.5) (p=0.05), respectively. CONCLUSION: ASXL1 and IDH mutations occur at a high incidence among young Egyptian AML patients with intermediate risk cytogenetics and confer a poorer outcome. Integration of mutations into risk profiling may predict outcome and impact therapeutic approach of young AML patient with uncertain prognosis.

Apoptosis and oxidative stress as relevant mechanisms of antitumor activity and genotoxicity of ZnO-NPs alone and in combination with N-acetyl cysteine in tumor-bearing mice.

Background: Several in vitro studies have revealed that zinc oxide nanoparticles (ZnO-NPs) were able to target cancerous cells selectively with minimal damage to healthy cells. Purpose: In the current study, we aimed to evaluate the antitumor activity of ZnO-NPs in Ehrlich solid carcinoma (ESC) bearing mice by measuring their effect on the expression levels of P53, Bax and Bcl2 genes as indicators of apoptotic induction in tumor tissues. Also, we assessed the potential ameliorative or potentiation effect of 100 mg/kg N-acetyl cysteine (NAC) in combination with ZnO-NPs. Materials and methods: ESC bearing mice were gavaged with three different doses of ZnO-NPs (50, 300 and 500 mg/kg body weight) alone or in combination with NAC for seven consecutive days. In addition to measuring the tumor size, pathological changes, zinc content, oxidative stress biomarkers and DNA damage in ESC, normal muscle, liver and kidney tissues were assessed. Results: Data revealed a significant reduction in tumor size with a significant increase in p53 and Bax and decrease in Bcl2 expression levels in the tissues of ZnO-NPs treated ESC bearing mice. Moreover, a significant elevation of MDA accompanied with a significant reduction of CAT and GST. Also, a marked increase in all comet assay parameters was detected in ZnO-NPs treated groups. On the other hand, the combined treatment with ZnO-NPs and NAC significantly reduced reactive oxygen species production and DNA damage in liver and kidney tissues in all ZnO-NPs treated groups. Conclusion: ZnO-NPs exhibited a promising anticancer efficacy in ESC, this could serve as a foundation for developing new cancer therapeutics. Meanwhile, the combined treatment with ZnO-NPs and NAC could act as a protective method for the healthy normal tissue against ZnO-NPs toxicity, without affecting its antitumor activity.

Role of mitochondria in rescuing glycolytically inhibited subpopulation of triple negative but not hormone-responsive breast cancer cells.

Triple-negative breast cancer (TNBC) subtype is among the most aggressive cancers with the worst prognosis and least therapeutic targetability while being more likely to spread and recur. Cancer transformations profoundly alter cellular metabolism by increasing glucose consumption via glycolysis to support tumorigenesis. Here we confirm that relative to ER-positive cells (MCF7), TNBC cells (MBA-MD-231) rely more on glycolysis thus providing a rationale to target these cells with glycolytic inhibitors. Indeed, iodoacetate (IA), an effective GAPDH inhibitor, caused about 70% drop in MDA-MB-231 cell viability at 20 µM while 40 µM IA was needed to decrease MCF7 cell viability only by 30% within 4 hours of treatment. However, the triple negative cells showed strong ability to recover after 24 h whereas MCF7 cells were completely eliminated at concentrations <10 µM. To understand the mechanism of MDA-MB-231 cell survival, we studied metabolic modulations associated with acute and extended treatment with IA. The resilient TNBC cell population showed a significantly greater count of cells with active mitochondria, lower apoptotic markers, normal cell cycle regulations, moderately lowered ROS, but increased mRNA levels of p27 and PARP1; all compatible with enhanced cell survival. Our results highlight an interplay between PARP and mitochondrial oxidative phosphorylation in TNBC that comes into play in response to glycolytic disruption. In the light of these findings, we suggest that combined treatment with PARP and mitochondrial inhibitors may provide novel therapeutic strategy against TNBC.

Bone marrow derived-mesenchymal stem cells downregulate IL17A dependent IL6/STAT3 signaling pathway in CCl4-induced rat liver fibrosis.

Therapeutic potential of bone marrow-derived mesenchymal stem cells (BM-MSCs) has been reported in several animal models of liver fibrosis. Interleukin (IL) 17A, IL6 and Stat3 have been described to play crucial roles in chronic liver injury. However, the modulatory effect of MSCs on these markers was controversial in different diseases. BM-MSCs might activate the IL6/STAT3 signaling pathway and promote cell invasion in hepatocellular carcinoma, but the immunomodulatory role of BM-MSCs on IL17A/IL6/STAT3 was not fully elucidated in liver fibrosis. In the present study, we evaluated the capacity of the BM-MSCs in the modulation of cytokines milieu and signal transducers, based on unique inflammatory genes Il17a and Il17f and their receptors Il17rc and their effect on the IL6/STAT3 pathway in CCl4-induced liver fibrosis in rats. A single dose of BM-MSCs was administered to the group with induced liver fibrosis, and the genes and proteins of interest were evaluated along six weeks after treatment. Our results showed a significant downregulation of Il17a, Il17ra, il17f and Il17rc genes. In accordance, BM-MSCs administration declined IL17, IL2 and IL6 serum proteins and downregulated IL17A and IL17RA proteins in liver tissue. Interestingly, BM-MSCs downregulated both Stat3 mRNA expression and p-STAT3, while Stat5a gene was downregulated and p-STAT5 protein was elevated. Also P-SMAD3 and TGFßR2 proteins were downregulated in response to BM-MSCs treatment. Collectively, we suggest that BM-MSCs might play an immunomodulatory role in the treatment of liver fibrosis through downregulation of IL17A affecting IL6/STAT3 signaling pathway.

Fibulin-2 is required for basement membrane integrity of mammary epithelium.

Fibulin-2 (FBLN2) is a secreted extracellular matrix glycoprotein which has been associated with tissue development and remodelling. In the mouse mammary gland, FBLN2 can be detected during ductal morphogenesis in cap cells and myoepithelial cells at puberty and early pregnancy, respectively. In an attempt to assign its function, we knocked down Fbln2 in the mouse mammary epithelial cell line EpH4. FBLN2 reduction led to an increase in the size of spheroidal structures when compared to scrambled control shRNA-transduced cells plated on Matrigel matrix. This phenotype was associated with a disruption of the collagen IV sheath around the epithelial spheroids and downregulation of integrin ß1, suggesting a role for FBLN2 in stabilizing the basement membrane (BM). In contrast to mice, in normal adult human breast tissue, FBLN2 was detected in ductal stroma, and in the interlobular stroma, but was not detectable within the lobular regions. In tissue sections of 65 breast cancers FBLN2 staining was lost around malignant cells with retained staining in the neighbouring histologically normal tissue margins. These results are consistent with a role of FBLN2 in mammary epithelial BM stability, and that its down-regulation in breast cancer is associated with loss of the BM and early invasion.

Antioxidant activity and apoptotic induction as mechanisms of action of Withania somnifera (Ashwagandha) against a hepatocellular carcinoma cell line.

Objective To evaluate the antioxidant and apoptotic inductive effects of Withania somnifera (Ashwagandha) leaf extract against a hepatocellular carcinoma cell line. Methods After treating HepG2cells with Ashwagandha water extract (ASH-WX; 6.25 mg/ml-100 mg/ml), cell proliferation was assessed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Antioxidant activities (total antioxidant, glutathione S-transferase and glutathione reductase), Fas-ligand level, tumour necrosis factor-α (TNF-α) level and caspase-3, -8, and -9 activities were measured. Molecular modelling assessed the binding-free energies of Ashwagandha in the cyclin D1 receptor. Results The MTT assay demonstrated increased cytotoxicity following treatment of HepG2 cells with ASH-WX compared with control untreated cells and theIC50was 5% (approximately 5.0 mg/ml). Antioxidant activities, Fas-ligand levels and caspase-3, -8 and -9 activities significantly increased, while TNF-α level significantly decreased following ASH-WX treatment compared with control untreated cells. Molecular docking analysis revealed a good prediction of binding between cyclin D1 and Ashwagandha. There was significant accumulation of ASH-WX-treated HepG2cells in the G0/G1 and G2/M phases compared with the control untreated cells. Conclusion Ashwagandha could be a powerful antioxidant and a promising anticancer agent against HCC.

Leptin is overexpressed in the tumor microenvironment of obese patients with estrogen receptor positive breast cancer.

The present study aimed to investigate the potential role of leptin in the progression of breast cancer and the associated cell proliferation signalling pathway(s). A total of 44 female patients diagnosed with breast cancer and 24 healthy donors from Ain Shams University Hospitals (Cairo, Egypt) were enrolled in the present study. The present study assessed leptin expression in breast cancer tissues at the gene and protein level using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry. The results demonstrate that the expression of leptin was significantly higher in tissue of breast cancer samples from obese patients than overweight and control samples (P<0.001). ELISA results indicated a significant increase (P<0.001) of leptin expression in obese patients. To investigate whether there is any difference in leptin expression between the peripheral and tumor microenvironment blood of patients with breast cancer, the concentration of leptin was assessed in plasma from both using ELISA assays. The results demonstrated a statistically significant increase in the level of leptin in plasma samples from the tumor microenvironment of obese patients with estrogen receptor positive (ER+) breast cancer, compared with peripheral plasma samples. Furthermore, the leptin gene was overexpressed in obese ER+ breast cancer tissue. RT-qPCR was also performed to assess the expression of genes involved in proliferation pathways including leptin receptor (LEPR), aromatase, mitogen activated protein kinase (MAPK) and signal transducer and activator of transcription-3 (STAT3). A positive association between leptin expression, LEPR, aromatase, MAPK and STAT3 was detected in tissue samples of patients with breast cancer. The current study concluded that leptin may enhance breast cancer progression by inducing the expression of JAK/STAT3, ERK1/2 and estrogen pathways in obese patients breast cancer.