In-vitro Modulation of mTOR-HIF-1α Axis by TLR7/8 Agonist (Resiquimod) in B-Chronic Lymphocytic Leukemia.

Targeting toll-like receptors (TLRs), via TLR agonists, has been implicated in the regulation of immunometabolism. B-chronic lymphocytic leukemia (B-CLL) represents a suitable model for B-cell derived malignancies with shifted metabolic adaptations. Several signaling pathways have been found to be critical in metabolic reprogramming of CLL, including mechanistic target of rapamycin- hypoxia inducible factor-1α (mTOR- HIF-1α) pathway, the main metabolic regulator of glycolysis. Here, we investigated the effect of TLR7/8 agonist (Resiquimod) on the expression of mTOR and HIF-1α in patients with CLL. B cells were purified using Rosettesep Human B cell Enrichment Cocktail (Stem cell Technologies, Vancouver, BC, Canada#15,024) from peripheral venous blood of CLL patients (n = 20) and healthy individuals (n = 15). Isolated B cells were then cultured in both presence and absence of Resiquimod. Gene expression of mTOR and HIF-1α were assessed using qRT-PCR. Resiquimod significantly decreased mTOR and HIF-1α gene expression in both CLL (p < 0.001and p < 0.001, respectively) and Normal B cells (p = 0.004 and p = 0.001, respectively). Resiquimod may reprogram immunometabolism of malignant B-CLL cells via down-regulation of key glycolytic metabolic actors, mTOR and HIF-1α genes. Accordingly, Resiquimod may be an adjuvant as a therapeutic tool for CLL, which needs to be studied further. Supplementary Information: The online version contains supplementary material available at 10.1007/s12288-023-01649-y.

IL-6 Impairs the Activity of Vitamin D3 in the Regulation of Epithelial-Mesenchymal Transition in Triple Negative Breast Cancer.

Objective: The present study aimed to investigate the possible role of IL-6 and 1α,25-dihydroxyvitamin D3 (1,25D) signaling in epithelial-mesenchymal transition (EMT) and stemness in triple-negative breast cancer (TNBC) cell line. Methods: TNBC cell line, HCC 1806, was treated with IL-6 and 1,25D for three and six days. Also, the role of vitamin D receptor (VDR) was studied by transfection of TNBC cell line with VDR gene and transfection efficiency was assessed using Human VDR enzyme-linked immunosorbent assay (ELISA). Changes in E-cadherin gene expression were analyzed by quantitative real-time PCR (qRT-PCR). Also, changes in CD44+ cells were analyzed by flow cytometry. Finally, morphological changes were investigated by light microscopy after 6 days. Results: Treatment of HCC1806 cells with IL-6 has no significant effect either on E-cadherin gene expression or CD44+ cells, (p > 0.05). However, E-cadherin gene expression was significantly up-regulated after treatment with 1,25D for 6 days, (p < 0.05). Also, CD44+ cells were significantly reduced after treatment with 1,25D either for 3 or 6 days, (p < 0.05). Transfection of TNBC cell line with VDR gene significantly up-regulated VDR protein expression, (p < 0.05). In addition, overexpression of VDR in TNBC cells and treatment with 1,25D significantly up-regulated E-cadherin gene expression, (p < 0.05) and reduced CD44+ cells, (p < 0.05). Moreover, transfection with VDR and treatment with a combination of 1,25D and IL-6 significantly down-regulated E-cadherin gene expression and increased CD44+ cells compared with transfected cells with VDR treated with 1,25D alone, (p < 0.05). No significant morphological changes were observed in treated cells, 6 days post-treatment. Conclusion: The presence of IL-6 in the breast tumor microenvironment may impair the activity of vitamin D3 signaling, limiting its anti-tumor effects in TNBC.

Changes of serum growth factors (IGF-I & IGFBP-2) and prediction of response to chemotherapy in patients with acute myeloid leukemia.

Insulin-like growth factors (IGFs) and IGF-binding proteins (IGFBPs) possess mitogenic properties promoting cellular proliferation and inhibiting cellular apoptosis. Thus, IGF-system may play a role in tumor proliferation. In the present study, we investigated IGF-I and IGFBP-2 in patients with acute myeloid leukemia (AML) for their predictive value to identify patients who could be responsive to conventional induction chemotherapy. Serum levels of IGF-I and IGFBP-2 were measured (using commercial ELISA kits) in 22 patients with AML. Following treatment, patients were followed up 14 days after the induction cycle to determine if they achieved hematological remission or not. Accordingly, patients were divided into two groups: Responders (10 patients) and Non-Responders (12 patients). No significant difference was observed in IGF-1 between Responders and Non-Responders, neither before (79.26 +/- 4.568 pg/ml vs 72.225 +/- 3.62 pg/ml, respectively) nor after induction cycle (74.87 +/- 3.669 pg/ml vs 69.783 +/- 4.329 pg/ml, respectively). However, IGFBP-2 was significantly lower in Responders than in Non-Responders both at diagnosis (4250 +/- 155.099 pg/ml vs 6866.67 +/- 352.122 pg/ml, respectively) as well as after induction cycle of chemotherapy (4130 +/- 324.225 pg/ml vs 7150.00 +/- 265.290 pg/ml, respectively). It is concluded that, serum IGFBP-2 is considered as an independent factor that adds additional information for the prediction of relapse or treatment failure and patients with concentration > or = 4900 pg/ml at diagnosis are suspected to be nonresponders to conventional induction chemotherapy.

Leukocytic-vascular endothelial growth factor and integrin alphavbeta3 in acute myeloid leukemia: relation to clinical outcome.

Different signaling routes seem to be simultaneously triggered in leukemia, with distinct and overlapping activities. Different reports emphasize the interaction between vascular endothelial growth factor (VEGF) and integrin alphavbeta3 as a key control system of angiogenesis, oncogenesis and metatasis. The current study was undertaken to investigate leukocytic-VEGF and integrin alphavbeta3 as correlated with clinical outcome in patients with acute myeloid leukemia (AML). The study groups included 10 newly diagnosed AML patients before the start of any chemotherapeutic medication and 10 normal healthy control subjects. The level of VEGF was estimated in culture supernatant of peripheral blood mononuclear cells (PBMN) of both groups using commercially available ELISA kit. The degree of integrin alphavbeta3 expression on PBMN was estimated by indirect immunoflourescence. Obtained results showed that the level of VEGF and degree of expression of integrin alphavbeta3 were significantly higher in AML patients than in normal healthy subjects. However, no significant correlation was observed between the levels of VEGF and the degree of expression of integrin alphavbeta3. When clinical findings were concerned, there was a significant positive correlation between VEGF and the percentage of blasts, both in peripheral blood & bone marrow. On the other hand, such correlations were not observed in case of integrin alphavbeta3. In addition no significant correlation was observed between either VEGF or integrin alphavbeta3 and clinical staging, age, and sex. In conclusion, our results proved the importance of VEGF and integrin alphavbeta3 in the pathogenesis of AML. However, the per se increased production or/and secretion of VEGF and integrin alphavbeta3 by leukemic PBMN cells, respectively can not be used as independent predictor (s) for clinical outcome in AML patients. It is more comprehensive to study changes of intracellular signaling pathways when such critically interacting factors are concerned in the leukemic process.

Influence of hepatitis C virus infection on circulating levels of sICAM-1 and VEGF in patients with hepatitis C and hepatocellular carcinoma (HCC) and their role in enhancing detection of HCC.

Infection with hepatitis C virus (HCV) is characterized by inflammatory liver damage and a long viral persistence associated with an increased risk of developing hepatocellular carcinoma (HCC). Intercellular adhesion molecule-1 (ICAM-1) plays a key role during liver inflammation and also expressed in HCC. Its cellular expression is associated with the release of soluble form (sICAM-1) in the peripheral blood. The process of angiogenesis plays a critical role in liver damage-associated HCV infection and in tumor growth and metastasis. Vascular Endothelial Growth Factor (VEGF) is an important angiogenic factor regulating tumor angiogenesis. This study aimed at investigating the influence of HCV infection on serum profile of sICAM-1 and VEGF in patients with hepatitis C and HCC and their diagnostic value as useful markers reflecting progressive liver damage and development of HCC. Serum levels of sICAM-1 and VEGF were determined in the serum of fifteen HCV infected patients, fifteen HCV-positive patients with superimposed HCC as well as ten healthy control subjects by enzyme linked immunosorbent assay. HCV RNA copy numbers were analyzed by Real-time polymerase chain reaction using TaqMan probe technology. Alpha-fetoprotein levels and serum aminotransferases activities were also measured. The group of patients with hepatitis C and superimposed HCC had significantly higher sICAM-1 and VEGF values than HCV infected patients (1178.113 +/- 631.87 vs. 313.67 +/- 82.72 & 320.88 +/- 117.99 vs. 132.45 +/- 91.56, p < 0.001 respectively). In comparison to healthy subjects, HCV infected patients showed dramatically elevated serum levels of VEGF (132.45 +/- 91.56 vs. 7.76 +/- 7.41, p < 0.001). On the other hand, sICAM-1 levels were elevated in patients with HCV as compared with healthy controls, but this did not reach statistical significance (313.67 +/- 82.72 vs. 230.3 +/- 47.4, p > 0.05). A highly significant correlation was found between VEGF and sICAM-1 levels in all patients (r = 0.731, p < 0.001) also between VEGF, sICAM-1 and AFP (r = 0.473, p < 0.001, r = 0.690, p < 0.001, respectively) as well as between sICAM-1 and AST activities (r = 0.367, p < 0.05). A weak correlation was found between the level of viremia and VEGF, sICAM-1 levels, yet this did not reach statistical significance (r = 0.312, p = 0.09 & r = 0.228, p > 0.05 respectively). The sensitivity of HCC detection using AFP alone was 93.3%. It yielded 100% detection sensitivity when combined with sICAM-1 and/or VEGF with diagnostic accuracy reaching 96.67%. In conclusion, HCV infection and the development of HCC on top greatly affect the serum profile of VEGF and sICAM-1. VEGF as it stimulates endothelial cell growth, it could modulate the expression of sICAM-1 and both could be considered as convenient markers of progressive liver damage, endothelial activation and therefore could improve detection and management of HCC.

Antibody profile, interferon-gamma and nutritional status in cryptosporidial infection among school children.

Cryptosporidium parvum is more common in children than in adults. The host responses that induce protective immunity against Cryptosporidium is poorly understood. The present work studied the antibody profile, interferon gamma (IFN-gamma) serum level and nutritional status among infected school children, aged eleven years in a rural school in Abis 8 village. Fecal examination showed nineteen cases only infected with Cryptosporidium and ten children free from parasites (controls). The IgA, IgG and IgM serum levels in infected cases were within normal level, without significant differences as compared to controls. The IgE serum level was significantly high in infected children. The cytokine IFN-gamma was significantly lower in the infected cases as compared to controls. Malnutrition was diagnostic among infected children. It was concluded that malnutrition is the important finding related to Cryptosporidium infection. Malnutrition decreased IFN-gamma the recognized factor in protection from infection and elevated serum IgE level.