Iranian Biomedical Journal最新文献

Introduction: The Warburg effect is considered one of the most important metabolic alterations in tumor cells. While extensively studied in solid tumors, the role of this effect in acute myeloid leukemia (AML) is less defined. hypoxia inducible factor 1 alpha (HIF-1α) and lactate dehydrogenase A (LDH-A) are key regulators of glycolysis, promoting lactate production and the expression of lactate transporters. In this study, we examined the impact of HIF-1α and LDH-A inhibition on metabolic pathways, cell viability, and lactate transporter mRNA expression in AML cell lines.

Methods: K-562 and HL-60 cells were treated with silibinin, an HIF-1α inhibitor, and sodium oxamate, a LDH-A inhibitor. Cell viability and apoptosis were evaluated using the MTT assay and flow cytometry, respectively. Acidification rate and LDH-A activity were assessed using lactate assay and LDH-A assay kits, respectively. Relative mRNA expression of MCT1 and MCT4 was determined using qRT-PCR.

Results: Treatment with silibinin and sodium oxamate reduced proliferation and increased apoptosis in both K-562 and HL-60 cells. As expected, both agents decreased extracellular lactate release in K-562 cells, and sodium oxamate inhibited the LDH-A activity in both cell lines. Interestingly, the expression of MCT1, but not MCT4, was downregulated in K-562 cells after treatment.

Conclusion: Our findings show that HIF-1α and LDH-A inhibitors not only serve as cytotoxic drugs but also regulate the expression of lactate transporter and interfere with the metabolism-related mechanisms in AML cells.

In ovarian cancer, ovarian cancer stem cells (OCSCs) not only contribute to tumor formation but also drive the progression, metastasis, and chemoresistance. The defining properties of OCSCs are sustained through the Wnt/β-catenin, Notch, and Hedgehog signaling pathways, which can synergize rather than act independently to form a triad that enhances key functional capabilities of OCSCs. Various therapeutic agents have been studied to target these pathways in OCSCs, including γ-secretase inhibitors (nirogacestat), smoothened inhibitors (vismodegib and sonidegib), and Wnt/β-catenin inhibitors (PRI-724 and ipafricept). While these agents have demonstrated antitumor activity in preclinical and early clinical studies, their clinical use remains challenging due to compensatory interactions among signaling pathways, which diminish the efficacy of single-agent treatments. To improve treatment outcomes, strategies involving combination approaches and personalized treatments need to be explored. This review aims to summarize current evidence on the role of Wnt/β-catenin, Notch, and Hedgehog signaling pathways in OCSCs and their therapeutic implications in ovarian cancer.

Background: Infertility is one of the major complications of diabetes. Diabetic-mediated oxidative stress could destroy autophagy in testes, leading to damaged sperm and male infertility. Stevia rebaudiana Bertoni is a medicinal herb with various health-beneficial effects, such as improving diabetes-induced infertility. However, till now, no exact mechanism is known for its effects on diabetic testes. This study aimed to investigate the anti-infertility role of S. rebaudiana aqueous extract in diabetic rats induced by streptozotocin (STZ) by focusing on autophagy and apoptosis pathways and antioxidant activity.

Methods: Male rats were divided into four groups of five, including a healthy control group, a diabetic control (STZ) group, and metformin (MET)-treated (500 mg/kg) and stevia-treated (400 mg/kg) diabetic groups for 30 days. After the experimental period was over, rats were sacrificed. Testes tissue and blood samples were prepared for biochemical, molecular, and histopathological analyses.

Results: Treatments with stevia and MET significantly increased body weight and testosterone levels and decreased FBS compared to the STZ group. Moreover, both treatments decreased the mRNA expression of Akt1, Mtor, Sqstm1/p62, Nfkb1 and caspase-9 and increased the expression of Becn1, Atg3, Atg5, Atg7, Map1lc3b/LC3B, Nrf2 and Bcl2, suggesting a possible improvement in autophagy- and apoptosis-related responses in diabetic rats. Histopathological changes and sperm characteristics also showed improvement following stevia and MET treatments.

Conclusions: Stevia aqueous extract could exert potential therapeutic effects on diabetes-induced infertility in diabetic rats, possibly through the modulation of autophagy, apoptosis, and antioxidant activity.

Background: Osteogenesis imperfecta is a rare hereditary disorder affecting bone and connective tissue. While most cases are linked to autosomal dominant mutations in the COL1A1 and COL1A2 genes, FKBP10 variants are associated with the autosomal recessive form of OI, type XI. The study represents the first cohort-based evaluation of the FKBP10 mutational spectrum in Iranian patients, leading to the discovery of a novel variant.

Methods: Thirty Iranian patients clinically diagnosed with OI were enrolled for genetic analysis. WES was performed to identify pathogenic variants, validated by PCR and sanger sequencing in patients and their parents. To explore the biological relevance of the identified variants, we constructed PPI networks and performed functional enrichment analysis using the ClueGO plugin. MD simulations with GROMACS were used to assess the structural impact of the mutations.

Results: Among 30 families, four exhibited pathogenic FKBP10 variants. Three patients were homozygous for the previously reported mutation (c.831dupC: p. G278Rfs95), while the fourth harbored a novel homozygous deletion (c.855_859del: p. G286Lfs84). Network analysis revealed significant involvement of CRTAP, IFITM5, SERPINF1, PPIB, FKBP10, P3H1, SERPINH1, and PLOD2 in collagen-related pathways. Computational modeling and MD simulations indicated reduced flexibility and more compact folding in the mutant FKBP10 protein, which aligns with impaired protein function and defective collagen processing.

Conclusion: This study reports a novel FKBP10 variant and presents the first cohort-based analysis of FKBP10 mutations in Iranian patients with OI. It demonstrates the value of combining WES with computational modeling to elucidate the molecular mechanisms underlying OI.

Background: Breast cancer (BC) is among the most prevalent malignancies in women worldwide, yet early diagnosis is associated with a high survival rate. The proliferation of BC is linked to the overexpression of genes within the PI3K/AKT/mTOR signaling pathway. piR-651 has been reported to be effective in the proliferation and metastasis of BC. This research sought to evaluate the impact of piR-651 inhibition on the PI3K/AKT/mTOR pathway in HUVEC, MCF-7, and MDA-MB-231 cells.

Methods: Anti-piR-651 and non-target sequences were introduced into HUVEC, MCF-7, and MDA-MB-231 BC cells by lipofectamine transfection. After 48 hours, total RNA was extracted, and qRT-PCR assessed the gene expression of PI3K, AKT, and mTOR.

Results: Anti-piR-651 treatment significantly increased PI3K, AKT, and mTOR gene expression in HUVECs (p < 0.001). In contrast, PI3K and mTOR expression decreased in MCF-7 and MDA-MB-231 cells (p < 0.001), while AKT expression remained unchanged in MDA-MB-231 cells (p > 0.05). Correlations between these genes varied by cell type, with significant associations observed at p < 0.05 or p < 0.01, depending on the group.

Conclusion: piR-651 inhibition causes AKT to behave independently of PI3K and mTOR, particularly in MCF-7 cells, suggesting limited gene therapy potential for estrogen receptor-positive BC. Preliminary data indicate that piR-651 inhibition may reduce BC cell proliferation through effects on PI3K and mTOR.

Background: Accurate detection of actionable epidermal growth factor receptor ( EGFR) mutations is essential for guiding targeted therapy in non-small cell lung cancer (NSCLC). Liquid biopsy approaches using circulating tumor DNA (ctDNA) and exosomal DNA (exoDNA) offer noninvasive alternatives for molecular profiling. This study evaluated the diagnostic performance of nested PCR combined with sanger sequencing for detecting common EGFR mutations (exon 19 deletions and the L858R point mutation) in plasma samples from Iranian NSCLC patients.

Methods: In this retrospective observational study, blood samples were collected from 30 NSCLC patients with confirmed EGFR mutations. ctDNA was extracted from plasma and analyzed using nested PCR followed by sanger sequencing. Specificity was assessed in 20 EGFR–wild type NSCLC patients serving as controls. Diagnostic performance was further evaluated in relation to clinicopathological factors.

Results: EGFR mutations were detected in plasma ctDNA in 63.3% of patients. Detection sensitivity was significantly associated with tumor stage but was independent of mutation subtype, age, sex, or smoking status. The assay showed high specificity, with no false positive results in control samples (95% CI: 83.9–100.0%). Although exoDNA analysis demonstrated a higher sensitivity than ctDNA (76.6% vs. 63.3%), this difference was not statistically significant. Notably, combined analysis of ctDNA and exoDNA increased overall detection sensitivity to 80%.

Conclusions: Nested PCR with sanger sequencing represents a reliable rule in strategy for EGFR mutation detection in plasma. Integrating ctDNA and exoDNA analyses substantially improves sensitivity and may enhance noninvasive molecular diagnostics in NSCLC.

Genetically engineered probiotics (GEPs) aim to address transient colonization and the intra- and inter-subject variability that limit conventional probiotics. These strains utilize Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas editing, programmable gene circuits, and biosensors in chassis such as E. coli Nissle 1917 and L. lactis. This narrative review summarizes the current engineering toolkits and standards (e.g., SEVA), chassis selection criteria, biocontainment strategies, and translational requirements under CMC/GMP frameworks and discusses regulatory considerations for clinical translation. Representative examples include IL-10-secreting Lactococcus lactis and phenylalanine-metabolizing strains for phenylketonuria (SYNB1618/SYNB1934), which illustrate pharmacodynamic target engagement and short-term preclinical safety. We outline clinical advancements in predefined pharmacodynamics, durability of function, monitoring shedding and horizontal gene transfer, and genomic-microbiome-informed patient stratification. Systems modeling approaches (Genome-Scale Metabolic Model/ Agent-Based Model) are discussed as tools to guide rational design. GEPs offer programmable “sense-and-respond” therapeutics, with successful clinical adoption depending on durable efficacy, long-term safety, and clearly defined regulatory pathways.

Background: The increasing prevalence of S. aureus, especially methicillin-resistant strains, poses a major healthcare threat due to limited therapies. To address this challenge, we engineered the chimeric endolysin ZAM-MSC as a potent antibiotic alternative, using domain-fusion strategies to enhance antibacterial activity. We designed ZAM-MSC by integrating the catalytic (M23) and cell wall-binding (SH3b) domains of lysostaphin with the catalytic domain (CHAP) from endolysin SAL-1. Structural optimization was performed using AlphaFold2 prediction, AutoDock Vina docking, and GROMACS simulations to evaluate domain interactions, protein stability, and binding dynamics.

Methods: The chimeric construct was cloned into pCold I, expressed in E. coli, and purified under solubility-optimized conditions. Purified ZAM-MSC, at a minimum concentration of 3 μg, reduced bacterial optical density within 15 minutes, demonstrating potent lytic activity. Thermal stability assays indicated that ZAM-MSC retained its enzymatic activity over 80-90% across 4-37 °C, with only a 10-20% decrease at 25-37 °C after 30 minutes. NaCl stability tests revealed maximal activity in the absence of NaCl, with gradual reduction in enzyme activity by increasing NaCl concentrations.

Results: Cytotoxicity analysis via MTT assay on L929 fibroblast cells showed cell viabilities of ~85-90% ± 5% at the highest enzyme concentrations tested, with no detectable cytotoxic effect compared to untreated controls. Hemolysis assays confirmed nearly 100% red blood cell integrity across all tested enzyme concentrations, supporting its biocompatibility with mammalian cells.

Conclusion: Our findings establish ZAM-MSC as a highly promising therapeutic candidate, combining computational precision with robust experimental validation.

Background: De novo pyrimidine biosynthesis is essential for the survival of all living organisms. DHODH catalyzes the fourth step in this pathway. Inhibition of DHODH induces pyrimidine depletion and effectively eradicates microorganisms like H. pylori, which lacks the pyrimidine salvage pathway. Herein, we expressed rHp-DHODH, characterized its enzymatic activity, kinetics, and stability, and subsequently evaluated its inhibition by HQNO.

Methods: The gene fragment encoding the rHp-DHODH protein was synthesized, subcloned, and expressed in soluble form. The recombinant protein was purified, its identity was confirmed, and its activity was measured using a colorimetric reduction assay. Kinetic parameters and the effects of pH, temperature, and incubation time on the enzymatic activity were investigated. The inhibitory effect of HQNO on rHp-DHODH was evaluated using the DCIP reduction assay. The MIC of HQNO against H. pylori was determined, and its bactericidal/bacteriostatic effect was assessed.

Results: Optimal soluble expression of rHp-DHODH was achieved in BL21(DE3)pLysS. The enzyme exhibited a specific activity of 5.7 U/mg. Maximum activity was observed at pH 8.0 in Tris-HCl buffer at 25 °C. The Km values were 39.75 µM for DHO and 5.37 µM for CoQ10, with a kcat of 3.82 s-¹. The IC50 of HQNO against the recombinant enzyme was determined as 1.75 μM. HQNO inhibited H. pylori growth with an MIC of 0.5-1.0 µg/mL, displaying concentration-dependent bacteriostatic to bactericidal effects.

Conclusions: The rHp-DHODH and its optimized enzymatic assay provide a reliable platform for screening candidate inhibitors, such as HQNO, advancing drug development efforts against H. pylori infection.

The use of medicinal plants by diabetic patients dates back to ancient times. In recent years, numerous reports have been published on the efficacy and safety of many medicinal plants in the treatment of diabetes through various mechanisms. This review highlights the up-to-date proposed mechanisms of action of the most common antidiabetic herbs used in Persian medicine, comprising Cinnamomum zeylanicum, Trigonella foenum-graecum, Urtica dioica, Nigella sativa, Citrullus colocynthis, Silybum marianum, Zingiber officinale, Punica granatum, Salvia officinalis, Vaccinium arctostaphylos, and Momordica charantia, with support from clinical and experimental studies. Clinical research has shown significant reductions in blood glucose levels and insulin resistance, as well as improvements in diabetes-related symptoms, including digestive disorders and lipid dysregulation, accompanied by negligible adverse effects. Continuing to study how these plants work and how effectively they treat diabetes is important for using these natural treatments in modern medicine, offering affordable and safe options for diabetes patients.