International Journal of Molecular and Cellular Medicine最新文献

Cancer is the second leading cause of death worldwide, surpassed only by cardiovascular diseases. This study investigated the anticancer effects of recombinant Clostridium α-toxin on breast cancer, both in vitro and in vivo. The entire coding sequence of a codon-optimized α-toxin was designed, cloned into the pET28a (+) vector, and expressed as recombinant α-toxin in Escherichia coli (E. coli) BL 21(DE3) cells transformed with the recombinant plasmid. The recombinant α-toxin was purified using Ni²⁺ affinity chromatography, and its accuracy and purity were confirmed through sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis. The anticancer effects of purified α-toxin were then assessed in vitro and animal models against MCF-7 breast cancer cells. Protein analysis confirmed the presence of a 48 kDa band corresponding to the recombinant α-toxin. Additionally, the IC₅₀ values of α-toxin against MCF-7 cells at 24, 48, and 72 h were 407.3±2.392 μg/mL, 287.3±5.411 μg/mL, and 258.1±4.671 μg/mL, respectively. In vivo, results demonstrated a significant reduction in mean cancer nodule size following α-toxin treatment (p<0.001). These findings suggest that α-toxin may serve as a promising candidate for breast cancer therapy.

Chronic spontaneous urticaria (CSU) is a skin disease caused by mast cells that produce inflammatory mediators. Immune checkpoint receptors such as program death-1 (PD-1) and T-cell immunoglobulin and mucin domain 3 (TIM-3) are essential for the pathophysiology of many autoimmune and allergic diseases. The aim of this study was to investigate the expression of PD-1 and TIM-3 in CSU patients and their relationship to the anti-inflammatory cytokines (TGF-β and IL-10). In the current study, peripheral blood mononuclear cells (PBMCs) from CSU patients and healthy individuals were used and the Urticaria Activity Score 7 (UAS7) was used to assess disease severity. TaqMan-based RT-PCR was used to assess the expression of TIM-3 and PD-1 as well as the anti-inflammatory cytokines transforming growth factor-β (TGF-β) and IL-10. The protein concentrations of TGF-β and IL-10 were also measured by ELISA. The relationship between the expression of TIM-3 and PD-1 as well as TGF- β and IL-10 and the severity of the disease was investigated. The results showed that PD-1 mRNA expression was significantly increased in CSU patients (P<0.0001), while TGF- β and IL-10 levels were higher in CSU patients, but this difference was not significant (p=0.638, p= 0.798). The increase in protein level of IL-10 was significant (P<0.0001). There was also a positive correlation between the expression of PD-1 and TGF- β molecules and disease activity (P=0.0043, P=0.0018). In conclusion, the study found that the immune system expresses inhibitory molecules and anti-inflammatory cytokines to control disease severity. The higher expression of PD-1 molecules and IL-10 is associated with disease severity, suggesting that the immune system is trying to control inflammation and reduce disease severity.

The role of memory T cells in orchestrating memory responses to previously known tumor antigens is well documented. The aim of this study was to assess the frequency of different memory T cell subsets in tumor-draining lymph nodes of patients with bladder cancer (BC) and their prognostic significance. Mononuclear cells were isolated from 50 tumor-draining lymph nodes of untreated patients with BC and stained with antibodies against the markers CD8, CD95, CD45RO and CCR7. Data were collected using the FACSCalibur flow cytometer and analyzed using FlowJo software. Among the CD8⁺ cytotoxic lymphocytes, the frequency of different subsets was determined including total memory cells (CD8⁺CD45RO⁺CD95⁺), T central memory (TCM: CD8⁺CCR7⁺CD45RO⁺CD95⁺), T effector memory (TEM: CD8⁺CCR7^-CD45RO⁺CD95⁺), T stem cell memory (TSCM: CD8⁺CCR7⁺CD45RO^-CD95⁺) and naïve T cells (CD8⁺CCR7⁺CD45RO^-CD95^-). The analysis revealed that on average 49.32±20.15 (between 1.62% and 87.20%) percent of CD8⁺ lymphocytes in draining lymph nodes of BC had a memory phenotype. TCM cells showed the highest frequency (34.71±17.04), while TSCM cells (7.51±8.53) demonstrated the lowest. The total frequency of memory cells tended to be higher in patients with tumor invasion to muscle layer (P=0.052) and stage III (P=0.042) than in patients without invasion and stage I. The TCM subset was more frequent in patients with necrotic tumors than in patients without necrosis (P=0.048). TSCM significantly increased in patients with N2 compared to N0 (P=0.042). Conversely, the ratio of TSCM cells to total memory cells was higher in lower tumor stages (P=0.059), tumors without muscle invasion (P=0.026) and low T grouping (P=0.043). Overall the data indicated an increase in the frequency of memory T cells and their TSCM and TCM cells with tumor progression. In contrast, the ratio of TSCM to total memory cells was higher in less advanced tumors. These results suggest that the immune system is frequently exposed to tumor antigens and strives to create a memory T cell reservoir, but this is suppressed by inhibitory factors provided by the tumor. These findings emphasize the importance of understanding the dynamic interplay between memory T cell subsets and BC progression.

Glioblastoma multiforme (GBM) is an aggressive cancer with a poor prognosis. Inflammation and angiogenesis are important processes in GBM that are interrelated. In this study, bioinformatic investigations were performed to detect common and key genes in the inflammatory and angiogenesis pathways of GBM. Additionally, relevant long non-coding RNAs (lncRNAs) were recognized as important gene regulators. Consequently, real-time PCR and correlation analyses were used to investigate changes in gene and lncRNA expression levels and explain their relationship. RELA emerged as a common key gene in these biological processes. LINC01366 and LINC01433 were identified as putative RELA regulators in different metabolic pathways using computational assays. According to our findings, the expression levels of RELA, LINC01366 and LINC01433 were found to be significantly upregulated in GBM samples. Correlational studies revealed a significant positive relationship of gene expressions between LINC01366 and LINC01433, indicating that they may have a coordinated effect on GBM biology. Nevertheless, there was no significant correlation between these lncRNAs and RELA. The current study highlights the high expression of LINC01366 and LINC01433 in GBM and emphasizes the importance of studying lncRNAs as putative regulators in the pathophysiology of GBM. Further research is needed to clarify their specific functions, in particular the associated inflammatory and angiogenesis pathways.

This research delves into the therapeutic implications of utilizing small interfering RNA (siRNA) to target the ribosomal protein S19 (RPS19) gene in chronic myeloid leukemia (CML) using the K562 cell line model. The primary objective was to investigate how gene silencing affects apoptosis promotion and cell cycle arrest. The study employed bioinformatics tools and databases to explore the interactions involving RPS19 and neighboring proteins. Subsequently, siRNA-mediated gene silencing was utilized to suppress RPS19 expression in K-562 cells, with assessments conducted on cell cycle progression and apoptosis through flow cytometry analysis. Furthermore, real-time PCR was employed to evaluate the expression levels of RPS19, along with the closely associated RPS16 and RPS18 genes. Silencing the RPS19 gene in siRNA-transfected K-562 cells led to an increase in apoptotic cells by over 20%, with a significant accumulation in the sub-G1 and G1 phases. Additionally, the knockdown of RPS19 resulted in a 75% decrease in RPS16 expression and a 50% decrease in RPS18 expression. These results demonstrate the therapeutic potential of targeting RPS19 in CML cells, suggesting a promising approach for precise treatment strategies in leukemia and potentially other types of cancer.

Hypoxia can cause significant changes in the glucose metabolism of cancer cells that prefer aerobic glycolysis for energy production instead of the conventional oxidative phosphorylation mechanism. In this study, breast cancer cells (MCF-7) were exposed to glucose (0-5.5-15-55 mM), during specific incubation periods (3, 6, 12, or 24 hours) under normoxic and hypoxic conditions. The expression levels of hypoxia-inducible factor-1α (HIF-1α), glucose transporter-1 (GLUT-1), and glycolytic enzymes at varying glucose concentrations in cells were investigated in the different oxygen environments. It was determined that glycolytic enzymes [Hexokinase 2 (HK2), Pyruvate Kinase M2 (PKM2), Glucose-6-phosphate dehydrogenase (G6PD), Lactate Dehydrogenase A (LDHA), Glyceraldehyde-3-phosphate Dehydrogenase (GAPDH), and Phosphofructokinase M (PFKM)] increased at the transcriptional level, especially in the first hours. This increase indicates that major metabolic reprogramming in response to hypoxia probably occurs over a short period of time. The increase in G6PD gene expression under high glucose and hypoxia conditions suggests that the pentose phosphate pathway (PPP) is used by cancer cells to synthesize necessary precursors for the cell. The results of the study showed that there is a significant interaction between hypoxia and glycolytic metabolism in cancer cells. It is thought that metabolic pathways activated by hypoxia and related genes located in these pathways will contribute to the literature by offering the potential to be target molecules for therapeutic purposes.

In a system biology-based study, we previously reported that IL-6 and IL6R -specific m-RNA levels were elevated in leukocytes of patients with Rheumatoid arthritis (RA). Here, the association of toll-like receptor4 (TLR4) rs 141534085 and cytochrome P450 family 51 subfamily A member 1(CYP51A1) rs6 with tumor necrosis factor-α (TNF- α), rheumatoid factor (RF)- and Anti- cyclic citrullinated peptide (anti-CCP) antibody -positivity was investigated in almost the same subjects. Forty-six patients and 48 normal subjects were recruited in this study. The blood leucocytes TLR4 rs 141534085 and CYP51A1 rs6 -comprising DNA sequences were amplified by using tetra-primer amplification refractory mutation system polymerase chain reaction (T-ARMS-PCR) technique and the PCR products were checked by Sanger DNA sequencing method. ELISA method was used to determine plasma levels of TNF- α, anti-CCP antibody and RF positivity of plasma was evaluated through a latex agglutination test. The TNF- α level was significantly higher in the patient group than control subjects (p= 0.001). Moreover, we were not able to find any correlation between TNF-α levels and RF as well as anti-CCP antibodies when we used the K²/ Fisher's exact test and Pearson test respectively. Our DNA sequencing data revealed the following new mutations in TLR4 rs141534085 comprising regions: A>T in position 1050, T>A in position 1052, and C>A in position 1054; and for CYP51A1 rs6 encompassing region, the new mutations were; G>A in position 21680, the T nucleotide was inserted in position 21762 and the G nucleotide was inserted in position 21763, G>T in position 21764. The data of this study showed that both TLR4 rs141534085 and CYP51A1 rs6 related DNA regions should be considered as hotspot areas in RA pathogenicity. Moreover, these data indicated that, TNF- α did not alter the production of anti-CCP and RF pathogenic antibodies in patients with long-term RA.

Liver cancer treatment faces significant obstacles such as resistance, recurrence, metastasis, and toxicity to healthy cells. Biometallic nanoparticles (NPs) have emerged as a promising approach to address these challenges. In this study, copper oxide-silver (Ag-doped CuO) NPs were prepared using a reduction method with Ephedra intermedia extract. The physicochemical properties of the NPs were evaluated using various techniques such as Field emission scanning electron microscopy (FESEM), Transmission Electron Microscope (TEM), X-ray diffraction (XRD), and Fourier-transform infrared spectroscopy (FTIR). Additionally, this study has evaluated nitric oxide levels (NO), reactive oxygen species (ROS) production, Bax, Bcl2, P53, and Caspase3 genes expression, as well as cell viability within 24 hours in liver cancer cell line HepG2. FESEM and TEM imaging confirmed the nanostructural nature of the synthesized particles with sizes ranging from 31.27 to 88.98 nanometers. XRD analysis confirmed the crystal structure of the NPs. Comparative analysis showed that the IC₅₀ values of the Ag-doped CuO NPs were significantly lower than that of the plant extracts. Molecular studies showed significantly increased expression of Bax, Caspase3, and P53 genes, inducing apoptosis in cancer cells, and downregulation of Bcl2 as a pro-metastasis gene. Additionally, the presence of Ag-doped CuO NPs significantly increased NO activity enzyme and ROS generation compared to the plant extract. The biosynthesized Ag-doped CuO NPs demonstrated the ability to induce apoptosis, increase ROS production, and enhance NO enzyme activity in HepG2 cancer cells, suggesting their potential as a therapeutic agent for liver cancer.

Esophageal cancer presents a challenge in gastroenterology and traditional chemotherapy and radiation therapy have less therapeutic activity with severe side effects. Thus, there is need for effective and safer alternatives. Probiotics, particularly Lactobacillus plantarum (L. plantarum) and its bacteriocins, might prevent or treat esophageal tumors. We aimed to investigate the use of L. plantarum and its bacteriocin as esophageal cancer therapy. First, we obtained 100 isolates of Lactobacillus spp. from dairy product samples. They screened for bacteriocin production and identified by PCR and gel electrophoresis for 16S ribosomal RNA gene. Bacteriocin was partially purified and tested against two different pathogens. Both L. plantarum and its bacteriocin were examined for cytotoxicity in vitro against esophageal cancer cell line (SK-GT4) and normal rat embryo fibroblast (REF) cells by MTT assay. Apoptosis was determined using an acridine orange /propidium iodide assay. The results showed that the isolate gives a high bacteriocin production about (2000AU/ml). In addition to antimicrobial activity, there was significant anticancer activity. L. plantarum had an IC₅₀ of 51.01 CFU/ml and bacteriocin IC₅₀ of 281.9 AU/ml against cancer cells. Both showed no cytotoxicity towards normal REF cells. Furthermore, there was a significant increase in apoptosis induction and in caspase-3 activity in cancer cells treated with L. plantarum and bacteriocin compared to untreated cells. In conclusion, L. plantarum and its bacteriocin show potent killing effect against esophageal cancer cells with no effect against normal cells indicating safety and selectivity with activation of apoptosis via caspase-3 induction suggesting potential clinical advantage.

Glioblastoma (GBM) is the most aggressive and lethal brain tumor. Artificial neural networks (ANNs) have the potential to make accurate predictions and improve decision making. The aim of this study was to create an ANN model to predict 15-month survival in GBM patients according to gene expression databases. Genomic data of GBM were downloaded from the CGGA, TCGA, MYO, and CPTAC. Logistic regression (LR) and ANN model were used. Age, gender, IDH wild-type/mutant and the 31 most important genes from our previous study, were determined as input factors for the established ANN model. 15-month survival time was used to evaluate the results. The normalized importance scores of each covariate were calculated using the selected ANN model. The area under a receiver operating characteristic (ROC) curve (AUC), Hosmer-Lemeshow (H-L) statistic and accuracy of prediction were measured to evaluate the two models. SPSS 26 was utilized. A total of 551 patients (61% male, mean age 55.5 ± 13.3 years) patients were divided into training, testing, and validation datasets of 441, 55 and 55 patients, respectively. The main candidate genes found were: FN1, ICAM1, MYD88, IL10, and CCL2 with the ANN model; and MMP9, MYD88, and CDK4 with LR model. The AUCs were 0.71 for the LR and 0.81 for the ANN analysis. Compared to the LR model, the ANN model showed better results: Accuracy rate, 83.3 %; H-L statistic, 6.5 %; and AUC, 0.81 % of patients. The findings show that ANNs can accurately predict the 15-month survival in GBM patients and contribute to precise medical treatment.