The role of neuron-like cell lines and primary neuron cell models in unraveling the complexity of neurodegenerative diseases: a comprehensive review.

Neurodegenerative diseases (NDs) are characterized by the progressive loss of neurons. As to developing effective therapeutic interventions, it is crucial to understand the underlying mechanisms of NDs. Cellular models have become invaluable tools for studying the complex pathogenesis of NDs, offering insights into disease mechanisms, determining potential therapeutic targets, and aiding in drug discovery. This review provides a comprehensive overview of various cellular models used in ND research, focusing on Alzheimer's disease, Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis. Cell lines, such as SH-SY5Y and PC12 cells, have emerged as valuable tools due to their ease of use, reproducibility, and scalability. Additionally, co-culture models, involving the growth of distinct cell types like neurons and astrocytes together, are highlighted for simulating brain interactions and microenvironment. While cell lines cannot fully replicate the complexity of the human brain, they provide a scalable method for examining important aspects of neurodegenerative diseases. Advancements in cell line technologies, including the incorporation of patient-specific genetic variants and improved co-culture models, hold promise for enhancing our understanding and expediting the development of effective treatments. Integrating multiple cellular models and advanced technologies offers the potential for significant progress in unraveling the intricacies of these debilitating diseases and improving patient outcomes.

The Hydroalcoholic Extract of Saffron Protects PC12 Cells against Aluminum-Induced Cell Death and Oxidative Stress in Vitro.

BACKGROUND: Aluminum (Al) exposure is among the environmental risk factors that may involve in the pathogenesis of neurodegenerative diseases. Oxidative stress has a critical role in the Al-induced toxicity. Saffron is a plant with potent radical scavenging and anti-oxidative properties. This investigation was designed to evaluate the possible protective effects of saffron extract (SE) on aluminum maltolate (Almal)-induced oxidative stress and apoptosis in PC12 cell line. METHODS: In this in vitro study, PC12 cells were divided into four groups including control, Almal (500 µM), Almal+SE (50 µg/ml), and Almal+SE (100 µg/ml). After 48 hours of treatment with Almal in the absence and presence of SE, cell viability and apoptosis were determined using MTT (3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide) assay and Annexin V flow cytometry, respectively. Catalase activity was determined as an index of oxidative stress. Statistical analyses were performed using one-way ANOVA (SPSS version 16.0). P<0.05 was accepted as a statistically significant difference between groups. RESULTS: Almal decreased the PC12 cells viability dose-dependently (IC50=500µM). Co-treatment of 50 and 100 µg/ml of SE with 500 µM of Al increased cell viability to 79% (P=0.04) and 86% (P=0.02) of the control group, respectively. Al also increased PC12 cells apoptosis and catalase activity to 37 and 2.7 folds of those of the control group (P<0.001 and =0.001respectively). 100 µg/ml of SE blunted the effects of Al on the increased cell apoptosis (P=0.02) and changes in the catalase activity (P=0.003). CONCLUSION: SE has protective effects against Al-induced apoptosis and oxidative stress and may possess therapeutic values in the treatment of Al-neurotoxicity.

NDRG2 Regulates the Expression of Genes Involved in Epithelial Mesenchymal Transition of Prostate Cancer Cells.

BACKGROUND: Metastasis is the main cause of prostate cancer (PCa) death. The inhibitory effect of N-myc downstream-regulated gene 2 (NDRG2) on the invasiveness properties of PCa cells has been demonstrated previously. However, its underlying mechanisms have not yet been investigated. The present study aimed to investigate the effects of NDRG2 overexpression on the expression of genes involved in epithelial-mesenchymal transition (EMT) including E-cadherin (E-CAD), α- and ß-catenins, Slug and Snail, transforming growth factor (TGF)-α and -ß, and vascular endothelial growth factor (VEGF). METHODS: In the present in vitro study, LNCaP cells were divided into three groups, namely NDRG2 group (transfected with PSES-pAdenoVator-PSA-NDRG2-IRES-GFP plasmid), mock group (transfected with mock plasmid), and control group (without transfection). The effect of NDRG2 overexpression on the migration and invasion of LNCaP cells were investigated using the transwell assay. Real-time PCR was used for the evaluation of gene expression. For the statistical analyses, one-way ANOVA, student t test or Mann-Whitney U test were applied using the SPSS software (version 15.0). P values <0.05 were considered statistically significant. RESULTS: The results demonstrated that the overexpression of NDRG2 reduced the invasion and migration of LNCaP cells compared to the control and mock groups (P<0.001). A decreased expression of TGF-ß (P=0.002), VEGF (P=0.014), Slug (P=0.005), and Snail (P=0.012); and an increased expression of E-CAD (P=0.009) were observed following NDRG2 overexpression in LNCaP cells. CONCLUSION: The results of the present study suggest that NDRG2 inhibits the invasiveness properties of LNCaP cells probably through changes in the expression of genes involved in EMT.

Serum ischemia modified albumin is a possible new marker of oxidative stress in phenylketonuria.

The role of oxidative stress in the pathogenesis of phenylketonuria (PKU)-associated disorders has been implicated. Ischemia modified albumin (IMA) is a modified form of serum albumin, which is produced under the conditions of oxidative stress. The aim of this study was to measure the serum level of IMA in the PKU patients and to investigate its ability in predicting the status of oxidative stress in these patients. Fifty treated-PKU patients and fifty age- and sex-matched healthy subjects were included in the study. The blood samples were obtained and the serum level of phenylalanine (Phe) was measured using reverse phase HPLC method. The levels of IMA, malondialdehyde (MDA), gamma-glutamyl transferase (GGT) activity, and uric acid (UA) were determined using colorimetric methods. The levels of serum Phe, IMA, and MDA were significantly higher (p < 0.001) and the level of UA (p < 0.05) was lower in the PKU patients compared to control group. Serum IMA level was positively correlated with MDA (r = 0.585, p < 0.001) and UA (r = 0.6, p < 0.001). An inverse relationship was observed between the serum level of IMA and Phe (r = - 0.410, p < 0. 01). Results of the present study suggest that serum IMA level could be used as a novel marker for the evaluation of oxidative stress in the PKU patients.

Effects of Two-by-Two Combination Therapy with Valproic Acid, Lithium Chloride, and Celecoxib on the Angiogenesis of the Chicken Chorioallantoic Membrane.

BACKGROUND: The synergistic effects of valproic acid (VPA), lithium (Li), and celecoxib (CX) have been shown in combination therapy against the proliferation and metastasis of numerous cancers. Angiogenesis plays a critical role in the pathogenesis of tumor growth and metastasis. The aim of the present study was to evaluate the antiangiogenic effects of VPA, lithium chloride (LiCl), and CX, alone or in 2-by-2 combinations, using the chicken chorioallantoic membrane (CAM) assay. METHODS: Fertilized chicken eggs were randomly divided into 10 groups: control, VPA (1.8 and 3.6 µmol/CAM), Li (0.15 and 0.60 µmol/CAM), CX (0.02 and 0.08 µmol/CAM), VPA+Li, VPA+CX, and CX+Li (n=10 per group). A window was made on the eggshells and the CAMs were exposed to a filter disk containing VPA, LiCl, and CX, alone or in 2-by-2 combinations. The control CAMs were treated with distilled water (vehicle). Three days after the treatment, the number of vessel branch points was counted in each CAM. The data were analyzed using SPSS, version 15.One-way ANOVA, followed by the Tukey tests, was used to compare the groups. A P<0.05 was considered a statistically significant difference between the groups. RESULTS: According to the results, all the tested drugs decreased the number of the vessel branch points in a dose-dependent manner compared to the control group (P<0.001). In addition, combinations of the drugs were more effective in decreasing angiogenesis than the use of each drug alone. CONCLUSION: These findings suggest that 2-by-2 combinations of VPA, CX, and LiCl can be considered an effective antiangiogenesis therapeutic modality.

Expression and purification of TAT-NDRG2 recombinant protein and evaluation of its anti-proliferative effect on LNCaP cell line.

N-myc downstream regulated gene2 (NDRG2) belongs to tumor suppressor protein family of NDRG. Anti-proliferative and anti-metastasis of NDRG2 overexpression has been demonstrated in a number of tumors. The aim of this study was to fuse the gene of Trans Activator of Transcription (TAT) protein transduction domain with NDRG2 gene and express and purify TAT-NDRG2 fusion protein in order to investigate the effects of TAT-NDRG2 protein on proliferation and apoptosis of LNCaP prostate carcinoma cell line. pET28a-TAT-NDRG2 and pET28a-NDRG2 plasmids were constructed and transformed into E. coli-BL21(DE3). TAT-NDRG2 and NDRG2 proteins were expressed in the bacteria, purified using affinity chromatography and verified using western blotting. The effects of TAT-NDRG2 and NDRG2 protein treatment on LNCaP cells proliferation and apoptosis were evaluated using MTT assay and AnnexinV, 7-AAD flow cytometry assay, respectively. Western blot analysis confirmed the expression and purification of TAT-NDRG2 and NDRG2 proteins. Treatment of LNCaP cells with TAT-NDRG2 protein increased cell death and induced apoptosis significantly (P < 0.05) compared to control and NDRG2 protein-treated cells. These results suggest that TAT-NDRG2 protein can be considered as a therapeutic modality for the treatment of tumors.

Inhibitory Effect of Bunium Persicum Hydroalcoholic Extract on Glucose-Induced Albumin Glycation, Oxidation, and Aggregation In Vitro.

BACKGROUND: Glucose-induced protein glycation has been implicated in the progression of diabetic complications and age-related diseases. The anti-glycation potential of polyphenol-rich plant extracts has been shown previously. Bunium Persicum has been demonstrated to possess a high level of polyphenols. The aim of current in vitro study was to determine the possible inhibitory effect of Bunium Persicum hydroalcoholic extract (BPE) on glucose-induced bovine serum albumin (BSA) glycation, oxidation, and aggregation. METHODS: Folin-Ciocalteu assay was used to measure the content of total phenolic compounds of BPE. To test the in vitro effect of BPE on the formation of glycated BSA, thiol group oxidation, and protein aggregation of BSA, various concentrations of BPE were incubated with BSA and glucose at 37 °C for 72 hr. Glycation, thiol group oxidation, and aggregation of BSA were then measured using thiobarbituric acid, 2, 4-dinitrophenylhydrazine, and Congo red colorimetric methods, respectively. Data were analyzed using the SPSS software (version 16.0). One-way ANOVA followed by Tukey's post hoc test was used to compare group means. P<0.05 was accepted as the statistically significant difference between groups. RESULTS: The results demonstrated that the content of total phenolics of BPE was 122.41 mg gallic acid equivalents per gram dried extract. BPE (10, 15, and 30 µg/ml) significantly inhibited the formation of GA in a concentration-dependent manner. BPE also significantly decreased the levels of thiol group oxidation and BSA aggregation. CONCLUSION: The results showed that BPE has anti-glycation and antioxidant properties and might have therapeutic potentials in the prevention of glycation-mediated diabetic complications.

The Effects of NDRG2 Overexpression on Cell Proliferation and Invasiveness of SW48 Colorectal Cancer Cell Line.

BACKGROUND: Colorectal cancer (CRC) is one of the most common causes of cancer-related death in the world. The expression of N-myc downstream-regulated gene 2 (NDRG2) is down-regulated in CRC. The aim of this study was to investigate the effect of NDRG2 overexpression on cell proliferation and invasive potential of SW48 cells. METHODS: SW48 cells were transfected with a plasmid overexpressing NDRG2. After stable transfection, the effect of NDRG2 overexpression on cell proliferation was evaluated by MTT assay. The effects of NDRG2 overexpression on cell migration, invasion and cell motility and matrix metalloproteinase 9 (MMP9) activities were also investigated using matrigel transwell assay, wound healing assay and gelatin zymography, respectively. RESULTS: MTT assay showed that overexpression of NDRG2 caused attenuation of SW48 cell proliferation. Transwell and wound healing assay revealed that NDRG2 overexpression led to inhibition of migration, invasion, and motility of SW48 cells. The overexpression of NDRG2 also reduced the activity of secreted MMP-9. CONCLUSIONS: The results of this study suggest that NDRG2 overexpression inhibits proliferation and invasive potential of SW48 cells, which likely occurs via suppression of MMP-9 activity.

G22A Polymorphism of Adenosine Deaminase and its Association with Biochemical Characteristics of Gestational Diabetes Mellitus in an Iranian Population.

Adenosine deaminase (ADA) is an important regulator of insulin action. The single nucleotide polymorphism (SNP) G22A in the ADA gene decreases enzymatic activity of ADA. The aim of this study was to investigate the relationship between the SNP G22A and blood glycemic control, insulin resistance, and obesity of gestational diabetes mellitus (GDM) patients in an Iranian population. SNP G22A was determined in women with GDM (N=70) and healthy pregnant women (control, N=70) using polymerase chain reaction-restriction fragment length polymorphism. Fasting plasma glucose (FPG), hemoglobin A1C (HbA1c), plasma insulin levels and plasma lipids were measured using commercial kits. Homeostasis model of assessment for insulin resistance (HOMA-IR) was calculated. The distribution of genotypes and alleles among GDM patients was similar to that of the control group. FPG and HbA1c were significantly higher in GDM patients with GG genotype compared with GDM patients with GA+AA genotype and non-GDM patients. The frequency of GG genotype was significantly higher in obese GDM patients compared to lean GDM patients. The SNP G22A in the ADA gene was not associated with the risk of GDM in our population. GG genotype was associated with poor glycemic control and obesity in GDM patients.

Intrathecal Amylin and Salmon Calcitonin Affect Formalin Induced c-Fos Expression in the Spinal Cord of Rats.

BACKGROUND: Amylin and Salmon Calcitonin belong to the calcitonin family of peptides and have high affinity binding sites in the rat spinal cord. The aim of this study was to characterize receptors for Amylin and Salmon Calcitonin functionally in the spinal cord of rats. We assessed the expression of c-Fos in response to intraplantar formalin in the lumbar regions of the spinal cord in conscious rats. METHODS: Amylin (0.05 nmoles) or Salmon Calcitonin (0.005 nmoles) was administered intrathecally (i.t.) 10 minutes before the start of the formalin test. Antagonists were injected intrathecally 10 minutes before the administration of either of the peptides. RESULTS: Two hours after formalin stimulation, rats pretreated intrathecally by either Amylin or Salmon Calcitonin, showed lower numbers of c-Fos immunoreactive nuclei in their lumbar spinal cord as compared to rats pretreated with saline. These effects were reversed upon co-administration of either of the Amylin antagonists AC187 or rat amylin8-37, but not rat α-CGRP8-37 (.) A few cells with c-Fos immunoreactivity were found in the lumbar spinal cord of rats two hours after i.t. injection of saline, Amylin and/or Salmon Calcitonin. However, Fos-like immunoreactivity was increased in the lumbar spinal cord two hours after i.t. treatment of either of the antagonists AC187 and rat amylin8-37,when compared to saline treated rats. CONCLUSION: Both Amylin and Salmon Calcitonin inhibit formalin induced c-Fos expression in the rat lumbar spinal cord when administered intrathecally. Effects of the two peptides were possibly produced by undefined receptors.

Comparison of neutrophil to lymphocyte ratio, platelet to lymphocyte ratio, and C-reactive protein in predicting osteomyelitis in patients with diabetic foot ulcer.

Introduction: Neutrophil/lymphocyte ratio (NLR) and platelet/ lymphocyte ratio (PLR) are readily available and inexpensive biomarkers that have received great attention for diagnosing type 2 diabetes(T2DM) complications. The objective of the present cross-sectional study was to compare diagnostic values of these biomarkers with C-reactive protein(CRP) in detecting diabetic foot ulcer (DFU) and osteomyelitis (OS) and discriminating between the degree of DFU according to Wagner's classification. Methods: A total of 217 individuals (42 healthy controls, 40 T2DM patients without DFU, and 135 patients with DFU) were enrolled. The DFU patients were classified according to Wagner's classification into grade 1, grade 2, and grade 3. Blood samples were obtained and various biochemical and hematological parameters including creatine, CRP, HbA1c, NLR, and PLR were measured. Results: The levels of CRP, PLR, and NLR were significantly higher in the patients with DFU and OS compared to healthy controls and T2DM patients without DFU. The median values of CRP were correlated with the severity of DFU and increased with DFU grades. The highest values of CRP, NLR, and PLR were observed in the DFU patients with OS which were significantly higher than those of DFU patients with grades 1 and 2 as well as T2DM patients without DFU. The PLR and NLR had no significant performance in diagnosing DFU patients with grades 1 and 2 from the patients without DFU. Conclusion: NLR and PLR could be useful for diagnosing OS but cannot be used for detecting lower grades of DFU. CRP showed higher performance in detecting OS compared to PLR and NLR. Supplementary Information: The online version contains supplementary material available at 10.1007/s40200-023-01327-w.

The role of systemic inflammatory indices in predicting atrial fibrillation and its complications: a narrative review.

Atrial fibrillation (AF) is associated with increased morbidity and mortality. Inflammation and oxidative stress play critical roles in AF occurrence and its complications. Therefore, evaluating the circulating levels of inflammatory and oxidative stress biomarkers and their possible applications in AF diagnosis and management have been the focus of many efforts. The monocyte-to-high-density lipoprotein cholesterol ratio (MHR) and neutrophil-to-lymphocyte ratio (NLR) are two non-invasive, available, and established markers that serve as indicators of inflammation and oxidative stress. This review summarizes the current literature regarding alterations in the NLR, MHR, and other composite markers of systemic inflammation in AF patients. Moreover, this review discusses the clinical performance of these markers in predicting AF occurrence, recurrence, and disease outcomes. The PubMed, Scopus, and ScienceDirect online databases were searched for relevant studies using appropriate keywords, including "atrial fibrillation", "monocyte to high-density lipoprotein cholesterol ratio", and "neutrophil to lymphocyte ratio". The results of this review revealed the association of elevated levels of systemic inflammatory markers, specifically the NLR and MHR with AF and its complications. This finding indicates the potential role of subclinical inflammation in the development of AF, emphasizing its consideration in both the prevention and treatment of AF and associated complications. Despite these promising findings, the utilization of these markers in routine clinical settings faces challenges, including low specificity and sensitivity and varying cut-off values across different studies.

Annexin-A5 Overexpression Increases Sensitivity of MCF-7 and MCF-7/ADR Cells to Epirubicin.

Background: Multi-drug resistance is an important challenge in the chemotherapy of cancer. The role of annexin A5 (ANXA5) in the biology of cancer has been the focus of many studies. Breast Cancer (BC) is frequent cancer in women with high morbidity and mortality rate. The present study aimed to investigate the effects of ANXA5 overexpression on the anti-tumor activity of Epirubicin (EPI) in MCF-7 and MCF-7/ADR cells. Methods: MCF-7 and MCF-7/ADR cells were transfected with the pAdenoVator-CMV-ANXA5-IRES-GFP plasmid or mock plasmid. The overexpression of ANXA5 was evaluated using qPCR. The effects of ANXA5 overexpression and EPI on the cell viability of MCF-7 and MCF-7/ADR cells were measured using an MTT assay. Cell apoptosis was measured by annexin V/7-AAD flow cytometry assay. Results: Following the overexpression of ANXA5, the viability of MCF-7 and MCF-7/ADR was significantly decreased. Furthermore, the overexpression of ANXA5 in MCF-7 cells increased the cytotoxic effects of EPI in all doses and reduced the IC50 of EPI from 17.69 µM to 4.07 µM. Similarly, the overexpression of ANXA5 in MCF7-ADR cells reduced the IC50 of EPI from 27.3 µM to 6.69 µM. ANXA5 overexpression alone or combined with EPI treatment increased the apoptosis of MCF7 and MCF7-ADR cells. Conclusion: The results of the present study demonstrate that ANXA5 overexpression increases the sensitivity of MCF-7 and MCF-7/ADR to EPI, suggesting a possible beneficial role of ANXA5 in the therapy of BC.

Protective effects of Panax Ginseng against 131I-induced genotoxicity in patients with differentiated thyroid cancer.

BACKGROUND: Radioiodine (131I) therapy (RAIT) is associated with oxidative stress (OS)-induced DNA damage in patients with differentiated thyroid cancer (DTC). The goal of this study was to evaluate the possible ameliorating effects of Panax Ginseng (PG) on RAIT-induced genotoxicity in patients with DTC. MATERIALS AND METHODS: Forty DTC patients who had received 131I (100 to 175 mCi) were enrolled in this study. The patients were randomly classified (n = 10) into control, placebo, PG1 groups (receiving 500 mg/day of PG for 2 days before RAIT), and PG2 group (receiving 500 mg/day of PG for 2 days before to 1 day after RAIT). Blood samples were collected before and 2 days after RAIT. Lymphocyte micronuclei (MN) frequency was measured using the MN assay. Serum total antioxidant capacity (TAC) and ischemia-modified albumin (IMA) were measured using colorimetric assays. Serum albumin, blood urea nitrogen (BUN), creatinine, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) were measured using commercial kits. RESULTS: The mean of baseline MN frequency was the same in the four groups. RAIT increased the MN frequencies to at least three times the baseline values in the control (39 ± 5) and placebo groups (38 ± 6) (P < 0.001). PG caused a significant decrease in the MN frequencies in the treated groups compared to the control and placebo groups (P < 0.001). RAIT and PG administration had no significant effects on the serum IMA, TAC, and markers of liver and kidney toxicity. CONCLUSION: PG could be considered a useful remedy for the protection against RAIT-induced chromosomal damage in DCT patients.

Combined Effects of Annexin A5 Overexpression, 5-Fluorouracil Treatment, and Irradiation on Cell Viability of Caski Cervical Cancer Cell Line.

Cervical cancer is the fourth leading cause of cancer deaths in women globally. Combining gene therapy with chemo- and radiotherapy may improve cervical cancer treatment outcomes. This study evaluated the effects of Annexin A5(ANXA5) overexpression alongside 5-fluorouracil (5-FU) and irradiation on the viability of CaSki cervical squamous cell carcinoma (SCC) cells. pAdenoVator-CMV-ANXA5-IRES-GFP-plasmid and mock plasmid were transfected into CaSki cells using calcium-phosphate. Seventy-two hours post-transfection, GFP expression was quantified by fluorescence microscopy and flow cytometry to evaluate transfection efficiency. ANXA5 overexpression was confirmed via qPCR. Twenty-four hours post-transfection, cells received a single dose of 8 Gy and were treated with 1 and 2 µg/ml of 5-FU (IC50 = 2.783 µg/ml). Cell viability, apoptosis, cell cycle stage, and Bcl-2 and Bax gene expression were assessed via MTT, annexin V/7-AAD, PI staining, and qPCR assays, respectively. ANXA5 was overexpressed 31.5-fold compared to control (p < 0.0001). MTT assays showed ANXA5 overexpression dose-dependently reduced CaSki cell viability (p < 0.001). IC50 of 5-FU was reduced from 2.783 µg/mL to 1.794 µg/mL when combined with ANXA5 overexpression. Additive effects on cell death were observed for ANXA5 plus 5-FU or irradiation versus ANXA5 alone. Apoptosis assays indicated combinatorial treatment increased CaSki cell apoptosis over ANXA5 alone. Cell cycle analysis revealed ANXA5 arrested cell cycle at G1/S phases; the percentage of cells in the S phase further rose with combination treatment. Finally, combination therapy significantly decreased Bcl-2 expression and increased Bax versus control (p < 0.001). Altogether, ANXA5 overexpression alongside 5-FU and irradiation may improve cervical squamous cell carcinoma (SCC) treatment efficacy. Further, in vivo investigations are warranted to confirm these in vitro results.

Diagnostic values of ischemia modified albumin in diabetes-related complications: a narrative review.

Purpose: Oxidative stress (OS) plays a critical role in the onset and progression of macro and microvascular complications of diabetes mellitus (DM). Ischemia-modified albumin (IMA) is a novel and simple test for evaluating OS. In the present study, we reviewed the available information on the alteration of circulating IMA in DM and its possible prognostic and diagnostic value in DM-related complications. Methods: Relevant studies regarding IMA alteration in DM published until May 30, 2022 were extracted from Google Scholar, PubMed, Scopus, and Science Direct databases. The following key words were used: IMA, DM, diabetes complications, retinopathy, nephropathy, diabetic foot, and vascular complications. Results: This review revealed increased circulating IMA levels in the patients with type 1, type 2, and gestational DM. Furthermore, IMA showed a close relationship with the severity of DM complications including hyperglycemia, dyslipidemia, diabetic retinopathy, diabetic nephropathy, peripheral arterial disease, and diabetic foot ulcer. However, lack of assay standardization and low specificity are major obstacles to the use of IMA as a promising biomarker. Conclusion: IMA levels are associated with DM complications and can be applied as a practical test for evaluating the risk and predicting the severity of DM complications.

Targeted Overexpression of NDRG2 using Survivin Promoter Reduces Viability and Invasiveness of A549 Cell Line.

Background: Anti-tumor effects of N-myc Downstream Regulated Gene2 (NDRG2) have been demonstrated in many tumors. In the present study, NDRG2 was specifically overexpressed in lung cancer cell line using Survivin Promoter (Sur-P). Then, the effects of NDRG2 overexpression on viability, apoptosis, migration, and invasion of A549 cells were evaluated. Methods: Recombinant pAdenoVator-Sur-P-NDRG2-IRES-GFP plasmid harboring NDRG2 gene under transcriptional control of Sur-P and mock plasmid were constructed. A549 lung tumor cells and LX-2 cells (non-tumor cell line) were transfected with pAdenoVator-Sur-P-NDRG2-IRES-GFP, pAdenoVator-CMV-NDRG2-IRES-GFP, or mock plasmids. Tumor specificity of Sur-P was evaluated using fluorescent microscopy for GFP expression. The effects of NDRG2 overexpression on cell viability, apoptosis, and migration of A549 cells were measured using MTT, annexinV/7-AAD flow cytometry, and transwell migration assay, respectively. NDRG2 and matrix metalloproteinase-2 (MMP-2) expression were measured using real time-PCR. Results: pAdenoVator-Sur-P-NDRG2-IRES-GFP transfection resulted in a huge GFP expression in A549 cells, but not in LX-2 cells. The results of real time-PCR analysis also showed that pAdenoVator-Sur-P-NDRG2-IRES-GFP transfection led to an abundant NDRG2 expression in A549 cells. NDRG2 overexpression decreased A549 cell viability through increasing cell apoptosis. In addition, migration, invasion, and MMP-2 expression decreased following NDRG2 overexpression in A549 cells. Conclusion: The findings indicate that the targeted overexpression of NDRG2 using Sur-P can reduce the viability and invasiveness of A549 cells, suggesting possible benefits of this approach in lung cancer therapy.

The effect of Annexin A5 overexpression on invasiveness and expression of the genes involved in epithelial-mesenchymal transition of HCT 116 cell line.

Epithelial-to-mesenchymal transition (EMT) plays a critical role in colorectal cancer (CRC) metastasis. In the present study, we evaluated the effects of annexin A5 (ANXA5) overexpression on invasiveness as well as the expression of genes involved in EMT of HCT 116 cell line. PCMV6-AC-IRES-GFP plasmid harboring ANXA5 cDNA was constructed. HCT 116 cell line was transfected with recombinant plasmids using Lipofectamine 3000. Fluorescent microscopy was used to determine the efficiency of plasmid transfection. Cell viability was determined using the MTT assay. HCT 116 cell migration was evaluated using wound healing assay and transwell migration assay. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to measure the expression of genes involved in EMT. The results of RT-qPCR showed overexpression of ANXA5 compared to the control group. ANXA5 overexpression had no significant effects on cell viability but significantly decreased the rate of wound closure in the wound healing assay as well as the number of migrated cells in transwell assay. Furthermore, ANXA5 overexpression decreased the expression of N-cadherin, Snail, Slug, MMP-2, and MMP-9 while the expression of E-cadherin increased following ANXA5 overexpression. However, VEGF expression did not significantly change after ANXA5 overexpression. Results of the present study suggest that ANXA5 overexpression might have inhibitory effects on the metastasis of CRC through modulating the expression of EMT- related genes.

Association of serum kynurenine/tryptophan ratio with poor glycemic control in patients with type2 diabetes.

PURPOSE: The role of indoleamine 2,3-dioxygenase (IDO) has been shown in insulin resistance and metabolic syndrome. The present study aimed to measure serum IDO activity in patients with type 2 diabetes (T2DM) and to determine its association with glycemic control, oxidative stress, and insulin resistance. METHODS: Seventy-four patients with T2DM and 74 healthy subjects were selected to participate in this study. Fasting serum biochemical parameters including fasting blood sugar (FBS), HbA1c, insulin, uric acid, albumin, tryptophan, kynurenine, and total antioxidant capacity (TAC) were measured. HOMA-IR, QUICKI, and HOMA-B were calculated using serum FBS and insulin values. IDO activity was estimated using kynurenine/tryptophan ratio (KTR). Data were analyzed using SPSS software (Version 15) and p < 0.05 was considered as a significant difference. RESULTS: The findings showed higher levels of FBS, HbA1c, HOMA-IR, and KTR in the patients compared to the controls. TAC and HOMA-B were significantly lowered in the T2DM patients compared to controls. KTR was significantly correlated with the level of HbA1c, and T2DM patients with poor glycemic control (HbA1c ≤ 8) had significantly higher level of KTR. HOMA-B was significantly correlated with serum tryptophan and inversely correlated with HbA1c. CONCLUSION: Serum KTR is increased in T2DM patients with poor glycemic control. Potential clinical implications and possible pathogenic roles of IDO in T2DM development should be further elucidated.

Adrenomedullin increases cAMP accumulation and BDNF expression in rat DRG and spinal motor neurons.

OBJECTIVES: Adrenomedullin (AM) has high expression in the spinal cord. In this study, we investigated the expression of AM and its receptor components, including calcitonin receptor-like receptor (CLR) and receptor activity modifying proteins (RAMPs) in dorsal root ganglion (DRG) and spinal motor (SM) neurons. Furthermore, the effects of AM on cAMP/cAMP response element-binding protein (CREB), AKT/glycogen synthase kinase-3 beta (GSK-3ß) signaling pathways, and expressions of brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) were evaluated. MATERIALS AND METHODS: Rat embryonic DRG and SM neurons were isolated, purified, and cultured. Real-time PCR was used to assess expressions of AM, CLR, and RAMPs. cAMP levels, p-CREB, BDNF, and NT-3 were determined using an enzyme-linked immunosorbent assay. p-AKT and p-GSK-3ß levels were determined by western blotting. Real-time PCR showed expressions of AM, CLR, RAMP2, and RAMP3 in both DRG and SM neurons. RESULTS: AM increased cAMP accumulation and p-CREB levels in DRG and SM neurons. AM increased p-AKT and p-GSK-3ß in DRG, but not SM neurons. AM significantly increased BDNF expression in both DRG and SM neurons. There was also an increase in NT-3 level in both DRG and SM neurons, which is statistically significant in SM neurons. CONCLUSION: These results showed both DRG and SM neurons are targets of AM actions in the spinal cord. An increase in BDNF expression by AM in both DRG and SM neurons suggests the possible beneficial role of AM in protecting, survival, and regeneration of sensory and motor neurons.