Factor VII R353Q (rs6046), FGA A6534G (rs6050), and FGG C10034T (rs2066865) Gene Polymorphisms and Risk of Recurrent Pregnancy Loss in Iranian Women.

Recurrent pregnancy loss is a multi factorial and heterogeneous disorder defined as two or more consecutive pregnancy losses before 20 weeks' gestation. Gene polymorphisms including factor VII R353Q (rs6046), fibrinogen alpha chain A6534G (rs6050) and fibrinogen gamma chain C10034T (rs2066865) have potential role in thrombophilia and the relation between these three polymorphisms and an increased risk of venous thrombosis have been reported. As thrombophilia is associated with a considerable proportion of pregnancy loss and the association between these gene polymorphisms and recurrent pregnancy loss remains controversial, the aim of the present study was to evaluate the relation of these polymorphisms and recurrent pregnancy loss in Iranian women. A total of 144 women with a history of two or more consecutive miscarriages as the patient group and 150 healthy women with two live births and no history of pregnancy loss as the control group were included in the study. Polymerase chain reaction and restriction fragment length polymorphism were used for genotyping. The results were validated by DNA sequencing. The SPSS, SNPStats and Finch TV were used to analyze the results. Factor VII R353Q (rs6046) gene polymorphism showed a significant difference between RPL patients and the control group according to multiple logistic regression models [codominant (OR=0.38; 95% CI=0.23-0.63, P≤0.0001), dominant (OR=0.32; 95% CI=0.20-0.52, P≤0.0001), over dominant (OR=0.46; 95% CI=0.29-0.75, P=0.0017) and log-additive (OR=0.35; 95% CI=0.23-0.53, P≤0.0001)]. Fibrinogen alpha chain A6534G (rs6050) and fibrinogen gamma chain C10034T (rs2066865) gene polymorphisms showed no correlation with recurrent pregnancy loss. Factor VII R353Q (rs6046) gene polymorphism can be considered a risk factor for recurrent pregnancy loss. Further studies in larger populations are needed to confirm the findings.

CCN3/NOV Serum Levels in Non-Alcoholic Fatty Liver Disease (NAFLD) Patients in Comparison with the Healthy Group and their Correlation with TNF-α and IL-6.

INTRODUCTION: Adipokine irregularity leads to inflammation, endothelial dysfunction, insulin resistance (IR), and Non-Alcoholic Fatty Liver Disease (NAFLD). Previous studies linked NOV/CCN3 to obesity, IR, and inflammation, but no research has explored the connection between CCN3 serum levels and NAFLD. METHODS: This case-control study assessed CCN3, IL-6, adiponectin, and TNF-α serum levels in 80 NAFLD patients and 80 controls using ELISA kits. Biochemical parameters were measured with commercial kits and an auto analyzer. RESULTS: NAFLD patients exhibited significantly higher CCN3 (2399.85 ± 744.53 vs. 1712.84 ± 478.19 ng/ml), TNF-α, and IL-6 levels, and lower adiponectin levels compared to controls (P<0.0001). In the NAFLD group, CCN3 showed positive correlations with FBG, insulin, HOMA-IR, and TNF-α. Binary logistic regression analysis revealed increased NAFLD risk in the adjusted model (OR [95% CI] = 1.220 [1.315 -1.131]). A CCN3 cut-off value of 1898.0050 pg/mL differentiated NAFLD patients from controls with 78.8% sensitivity and 73.2% specificity. CONCLUSION: It was found that elevated CCN3 serum levels directly correlate with NAFLD incidence and inflammation markers (IL-6 and TNF-α). CCN3 could serve as a potential biomarker for NAFLD, but further research is needed to validate this finding and assess its clinical utility.

SGLT2 inhibitors and AMPK: The road to cellular housekeeping?

Sodium-glucose co-transporter-2 (SGLT2) inhibitors, known as Gliflozins, are a class of Glucose-lowering drugs in adults with type 2 diabetes (T2D) that induce glucosuria by blocking SGLT2 co-transporters in the proximal tubules. Several lines of evidence suggest that SGLT2 inhibitors regulate multiple mechanisms associated with the regulation of varying cellular pathways. The 5'-adenosine monophosphate-activated protein kinase (AMPK) pathway plays an important role in metabolic homeostasis by influencing cellular processes. Recently, it has been shown that SGLT2 inhibitors can affect the AMPK pathway in differing physiological and pathological ways, resulting in kidney, intestinal, cardiovascular, and liver protective effects. Additionally, they have therapeutic effects on nonalcoholic fatty liver disease and diabetes mellitus-associated complications. In this review, we summarize the results of studies of AMPK-associated therapeutic effects of SGLT2 inhibitors in different organelle functions.

Fibrinogen Aα gene genotyping in patients with inherited afibrinogenemia deficiency; a novel mutation in Iranian afibrinogenemia patients.

BACKGROUND: Congenital fibrinogen deficiencies (CFD) are a group of rare bleeding disorders (RBD). Afibrinogenemia as a subclass of these disorders would occurs as a result of mutations in fibrinogen gene. Here in, the sequences of Aα chain of fibrinogen (FGA) in patients with inherited afibrinogenemia disorder in south-eastern of Iran were analysed. METHODS: The FGA gene exons were amplified using PCR method and the DNA sequences were analysed to study the mutations in Aα chain of Fibrinogen. RESULTS: Results showed that there was no large deletion in FGA gene. Although a frame shift mutation: c.196_197insT p.Ser66PhefsX10 in a patient and a novel mutation of IVS2-1G>A in two other patients were detected which were different from those detected in European population. CONCLUSION: Different mutations are responsible of afibrinogenemia deficiency which requires more relevant studies for confirmation. The type and distribution of mutations in fibrinogen gene in Iranian patients is significantly different with reported mutations in European patients.

Curcumin and targeting of molecular and metabolic pathways in multiple sclerosis.

Multiple sclerosis (MS) is a life-threading disease that poses a great threat to the human being lifestyle. Having said extensive research in the realm of underlying mechanisms and treatment procedures, no definite remedy has been found. Over the past decades, many medicines have been disclosed to alleviate the symptoms and marking of MS. Meanwhile, the substantial efficacy of herbal medicines including curcumin must be underscored. Accumulated documents demonstrated the fundamental role of curcumin in the induction of the various signaling pathways. According to evidence, curcumin can play a role in mitochondrial dysfunction and apoptosis, autophagy, and mitophagy. Also, by targeting the signaling pathways AMPK, PGC-1α/PPARγ, and PI3K/Akt/mTOR, curcumin interferes with the metabolism of MS. The anti-inflammatory, antioxidant, and immune regulatory effects of this herbal compound are involved in its effectiveness against MS. Thus, the present review indicates the molecular and metabolic pathways associated with curcumin's various pharmacological actions on MS, as well as setting into context the many investigations that have noted curcumin-mediated regulatory effects in MS.

Expression of Long Non-Coding RNA H19 in Acute Lymphoblastic Leukemia.

OObjective: Long non-coding RNA (lncRNA) H19 has essential roles in growth, migration, invasion, and metastasis of most cancers. H19 dysregulation is present in a large number of solid tumors and leukemia. However, the expression level of H19 in acute lymphoblastic leukemia (ALL) has not been elucidated yet. The current study aimed to explore H19 expression in ALL patients and cell lines. MATERIALS AND METHODS: This experimental study was conducted in bone marrow (BM) samples collected from 25 patients with newly diagnosed ALL. In addition, we cultured the RPMI-8402, Jurkat, Ramos, and Daudi cell lines and assessed the effects of internal (hypoxia) and external (chemotherapy medications L-asparaginase [ASP] and vincristine [VCR]) factors on h19 expression. The expressions of H19, P53, c-Myc, HIF-1α and ß-actin were performed using quantitative real-time polymerase chain reaction (qRT-PCR) method. RESULTS: There was significantly increased H19 expression in the B-cell ALL (B-ALL, P<0.05), T-cell ALL (T-ALL, P<0.01) patients and the cell lines. This upregulation was governed by the P53, HIF-1α, and c-Myc transcription factors. We observed that increased c-Myc expression induced H19 expression; however, P53 adversely affected H19 expression. In addition, the results indicated that chemotherapy changed the gene expression pattern. There was a considerable decrease in H19 expression after exposure to chemotherapy medications; nonetheless, hypoxia induced H19 expression through P53 downregulation. CONCLUSION: Our findings suggest that H19 may have an important role in pathogenesis in ALL and may act as a promising and potential therapeutic target.

Gastrointestinal, Liver, Pancreas, Oral and Psychological Long-term Symptoms of COVID-19 After Recovery: A Review.

Due to the importance of control and prevention of COVID-19-correlated long-term symptoms, the present review article has summarized what has been currently known regarding the molecular and cellular mechanisms linking COVID-19 to important long-term complications including psychological complications, liver and gastrointestinal manifestations, oral signs as well as even diabetes. COVID-19 can directly affect the body cells through their Angiotensin-converting enzyme 2 (ACE-2) to induce inflammatory responses and cytokine storm. The cytokines cause the release of reactive oxygen species (ROS) and subsequently initiate and promote cell injuries. Another way, COVID-19-associated dysbiosis may be involved in GI pathogenesis. In addition, SARS-CoV-2 reduces butyrate-secreting bacteria and leads to the induction of hyperinflammation. Moreover, SARS-CoV-2-mediated endoplasmic reticulum stress induces de novo lipogenesis in hepatocytes, which leads to hepatic steatosis and inhibits autophagy via increasing mTOR. In pancreas tissue, the virus damages beta-cells and impairs insulin secretion. SARS-COV-2 may change the ACE2 activity by modifying ANGII levels in taste buds which leads to gustatory dysfunction. SARS-CoV-2 infection and its resulting stress can lead to severe inflammation that can subsequently alter neurotransmitter signals. This, in turn, negatively affects the structure of neurons and leads to mood and anxiety disorders. In conclusion, all the pathways mentioned earlier can play a crucial role in the disease's pathogenesis and related comorbidities. However, more studies are needed to clarify the underlying mechanism of the pathogenesis of the new coming virus.

Renal, cardiac, neurological, cutaneous and coagulopathic long-term manifestations of COVID-19 after recovery; A review.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused the novel global coronavirus disease 2019 (COVID-19) disease outbreak. Its pathogenesis is mostly located in the respiratory tract. However, other organs are also affected. Hence, realising how such a complex disturbance affects patients after recovery is crucial. Regarding the significance of control of COVID-19-related complications after recovery, the current study was designed to review the cellular and molecular mechanisms linking COVID-19 to significant long-term signs including renal and cardiac complications, cutaneous and neurological manifestations, as well as blood coagulation disorders. This virus can directly influence on the cells through Angiotensin converting enzyme 2 (ACE-2) to induce cytokine storm. Acute release of Interleukin-1 (IL1), IL6 and plasminogen activator inhibitor 1 (PAI-1) have been related to elevating risk of heart failure. Also, inflammatory cytokines like IL-8 and Tumour necrosis factor-α cause the secretion of von Willebrand factor (VWF) from human endothelial cells and then VWF binds to Neutrophil extracellular traps to induce thrombosis. On the other hand, the virus can damage the blood-brain barrier by increasing its permeability and subsequently enters into the central nervous system and the systemic circulation. Furthermore, SARS-induced ACE2-deficiency decreases [des-Arg9]-bradykinin (desArg9-BK) degradation in kidneys to induce inflammation, thrombotic problems, fibrosis and necrosis. Notably, the angiotensin II-angiotensin II type 1 receptor binding causes an increase in aldosterone and mineralocorticoid receptors on the surface of dendritic cells cells, leading to recalling macrophage and monocyte into inflammatory sites of skin. In conclusions, all the pathways play a key role in the pathogenesis of these disturbances. Nevertheless, more investigations are necessary to determine more pathogenetic mechanisms of the virus.

Bioactivity of Exosomes Derived from Trained Natural Killer Cells versus Non-Trained One: More Functional and Antitumor Activity.

Background: Natural killer (NK) cells are cytotoxic lymphocytes of the innate immune system, capable of killing viral-infected and cancerous cells. NK cell-mediated immunotherapy has remarkably changed the current paradigm of cancer treatment in recent years. It emerged as a safe and effective therapeutic approach for patients with advanced-stage leukemia. Several immune-escape mechanisms can be enacted by cancer cells to avoid NK-mediated killing. Exosomes released by NK cells that carry proteins and miRNAs can exert an antitumor effect. In the present study, we hypothesized that maybe exosomes derived from trained natural killer cells show more antitumor effect in comparison to non-trained one. Methods: PBMC was separated by the Ficoll method and cultured with IL-2 for 21 days to expand NK cells. The NK cells were co-cultured with K562 for 72 hours and exosome-derived co-cultured (as trained) and natural killer cell-derived exosomes (as non-trained) were extracted by Exo kit. The exosomes were confirmed by dynamic light scattering (DLS), transmission electron microscopy (TEM), flow cytometry, and western blotting. The K562 cells were separately treated by trained and non-trained exosomes and MTT assay, apoptosis, and real-time PCR were performed. Results: Based on flow cytometry, CD56 marker was 89.7% and 40.1% for NK cells and NK-derived exosomes, respectively. CD63 and CD9 were positive for exosomes by western blotting. The morphology of exosome was confirmed by TEM. Treated K562 cells by trained exosomes indicated the diminished cell viability and higher apoptosis. Furthermore, the trained exosomes showed up-regulation in both P53 and caspase3 genes as compared with non-trained sample. Discussion. Trained Exos showed a potent inhibitory effect on proliferation and induced apoptosis on K562 cell lines compared to the same dose of non-trained Exos. According to the results of qRT-PCR, trained Exos exerted an antitumor activity through up-regulation of caspase 3 and P53 in the apoptotic signaling pathway in tumor cells. Our findings indicate an effective action of trained Exos against cancer cells.

A Comparative Study of the Bone Marrow- and Umbilical Cord-Derived Mesenchymal Stem Cells (MSCs) Efficiency on Generating MSC-Educated Macrophages (MEMs).

BACKGROUND: Mesenchymal stem cells (MSCs) have gained much more attention in cell therapy and regenerative medicine due to their immunosuppressive effects. MSCs have interaction with other immune cells, such as macrophages (MQs). Bone marrow (BM)-derived MSCs can educate MQs toward MSC-educated MQs (MEMs) which possess an anti-inflammatory immunophenotype. Given this and based on the important limitations of BM collection, we hypothesized whether co-culture of MQs with umbilical cord (UC)-derived MSCs can result in the MEM phenotype. METHODS: First, isolated monocytes cultured for five days to obtain M0 MQs. Then, they were co-cultured with either BM- or UC-MSCs under direct and indirect conditions. After three days of co-culture, MEM-specific surface markers, as well as the gene expression of inflammatory and anti-inflammatory cytokines, were evaluated. RESULTS: Surface expression of CD163/CD206, as specific markers for M2 MQs, increased in MEMs after co-culture with BM- and UC-derived MSCs, while CD80/CD86 expression (specific markers for M1 MQs) didn't change significantly. The mRNA expressions of PDL-1 as well as anti-inflammatory cytokines, including IL-6, IL-10, and TGFß also increased in MEMs after co-culture of UC-MSCs compared to control MQs (p <.05), while the expression of IL-12 was significantly decreased (p<.001). CONCLUSIONS: To the best of our knowledge, this study shows for the first time that the co-culture of MQs with UC-derived MSCs efficiently contributes to the generation of MEMs even greater than BM-MSCs; shedding light on the promising potential of UC as an alternative source to educate MQs in vitro.

Hsa-miR-625 Upregulation Promotes Apoptosis in Acute Myeloid Leukemia Cell Line by Targeting Integrin-linked Kinase Pathway.

BACKGROUND: Growing evidence has demonstrated that microRNAs have a major effect on development of different types of cancer including AML. The overexpression of miR-625 could decrease tumorgenesis of acute myeloid leukemia cell lines through Integrin-linked kinase signaling pathway and reducing the associated oncogenes.  The aim of this study is to evaluate the effect of hsa-miR-625 upregulation on apoptosis and proliferation of KG1 cell line via ILK signaling pathway. METHODS: The KG-1 cell line was transfected with pLenti-III-premir625-GFP through viral method. Then, expression of miR-625 and genes were analyzed by quantitative PCR. Western blotting was used to evaluate for the protein level. Apoptosis was investigated by flow cytometry. Cell cycle analysis with PI and CCK-8 assay were performed to evaluate proliferation. RESULTS: KG-1 cells transfected with pLenti-III-pre mir625-GFP construct showed a significant increase in cell apoptosis. Gene expression of ILK and NF-κB were downregulated and AKT, c-fos, Caspase3, cyclin D1, KLF-4, OCT-4 and Nanog were upregulated but no alteration in GSK3 expression profile was observed. Downregulation of NF-κB and upregulation of Caspase 3 and p-ß-catenin protein levels were indicated (p<0.05). CONCLUSION: MiR-625 can be a promising approach to aid in the treatment of AML. However, further studies are required in this respect.

Expression and methylation status of vascular endothelial growth factor and thrombospondin-1 genes in congenital factor XIII-deficient patients with intracranial hemorrhage.

Congenital factor XIII (FXIII) deficiency is one of the rarest bleeding disorders, with an incidence of one per 2 million persons. Intracranial hemorrhage (ICH), a major cause of mortality in FXIII deficiency, is reported to be associated with vascular endothelial growth factor (VEGF) and thrombospondin-1 (TSP-1). Therefore, we investigated the association of VEGF and TSP-1 expression and methylation patterns with ICH in congenital FXIII deficiency patients. This study was conducted on 40 participants with FXIII, 20 of whom experienced ICH (cases), and 20 who did not (controls). Methylation pattern, gene expression, and plasma protein level were assessed using bisulfite sequencing PCR, quantitative real-time PCR, and ELISA. We found a partially methylated pattern for both VEGF and TSP-1 (P > 0.05). VEGF mRNA levels of the case group were significantly higher than those of the control group (P < 0.05), whereas TSP-1 mRNA levels did not show significant upregulation (P > 0.05). Plasma VEGF and TSP-1 concentrations in the case group were higher, but not statistically significant (P > 0.05). Our findings showed no obvious correlation between VEGF or TSP-1 methylation patterns and expression, suggesting that their expression in FXIII deficiency may not solely be controlled by gene methylation.

Promoter Methylation Status and Expression Levels of RASSF1A Gene in Different Phases of Acute Lymphoblastic Leukemia (ALL).

Background: Although the precise pathogenesis of acute lymphoblastic leukemia (ALL) remains unclear, studying gene-regulating mechanisms during ALL pathogeneses may shed light on the underlying mechanisms driving malignant behavior. There is some evidence showing the promoter hypermethylation and silencing of RASSF1A tumor suppressor gene in ALL cells; however, there is a lack of evidence for whether the gene indeed alters during different phases of ALL or in response to therapy. Thus, the current study aimed to clarify this issue using groups of adult ALL patients who have been scarcely investigated regarding expression levels and promoter methylation status. Materials and Methods: In this case/control study, the expression levels and methylation status of the gene promoter was evaluated using quantitative real-time PCR and methylation-specific PCR (MSP), respectively in adults with ALL. The study included peripheral blood of patients with newly diagnosed ALL (n=10), complete remission (CR) (n=10), or relapse (n=10), and 10 control samples from healthy individuals. Results: MSP results revealed an unmethylated status for almost all patients and control samples, except a case with relapsing ALL, which showed a hemimethylated pattern. RASSF1A also showed no difference in terms of gene expression in the patients compared with the control group (p>0.05). Conclusion: The results revealed an up-regulation of RASSF1A tumor suppressor in adult ALL patients experiencing CR, suggesting this to be a marker of therapy response. However, further investigations using more sensitive methylation detecting tools with larger sample sizes may better clarify the involvement of the promoter methylation of RASSF1A in these patients.

Validity of a Serological Diagnostic Kit for SARS-CoV-2 Available in Iran.

BACKGROUND: The Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) epidemic broke out in December 2019 and is now characterized as a pandemic. Effective control of this infectious disease requires access to diagnostic techniques, for both case finding and epidemic size estimation. The molecular technique is routinely used worldwide. Although it is the "standard" case detection and management method, it has its own shortcomings. Thus, some easy-to-use rapid serological tests have been developed. METHODS: One hundred and fourteen positive RT-PCR-diagnosed patients were tested by VivaDiag Kit, a brand of rapid serological kits available in hospitals affiliated to Tehran University of Medical Sciences (TUMS), Tehran, Iran. Frozen serum specimens taken from healthy people in summer and fall 2019 were also tested as negative controls. RESULTS: Test sensitivity was 47.9% (95% confidence interval [CI]: 38.8-56.9) for IgM and 47.0% (95% CI: 38.0-56.0) for IgG. There was no difference between IgG and IgM seropositivity except in one case. Specificity was calculated as 99.0% (95% CI: 96.4-99.9) for IgM and of 100.0% (95% CI: 0.98.2-100.0) for IgG. Sensitivity was higher in men and older participants. CONCLUSION: This test can be used for epidemiological investigations, especially for the estimation of the level of infection in the community, after it is properly corrected for sensitivity and specificity. The low sensitivity could be attributed to the technical limitations of the kit or low levels of antibodies after infection. The different sensitivity in age and sex groups supports the hypothesis that different people show different immune responses to this virus.

Comparison of Patient-collected and Lab Technician-collected Nasopharyngeal and Oropharyngeal Swabs for Detection of COVID-19 by RT-PCR.

BACKGROUND & OBJECTIVE: A simple approach to prevent close contact in healthcare settings during the COVID-19 outbreak is to train patients to collect their own nasopharyngeal and oropharyngeal swabs and deliver them to medical laboratories to have them processed. The aim of our study was to compare lab technician- with patient- collected oropharyngeal and nasopharyngeal samples for detection of the coronavirus disease 2019 (COVID 19) using rapid real-time polymerase chain reaction (rRT-PCR). METHODS: Fifty adult patients with flu-like symptoms and radiologic findings compatible with atypical pneumonia who were admitted to the infectious diseases ward of Imam Khomeini Hospital Complex, Tehran, Iran, with a clinical diagnosis of COVID-19 from February 28 to April 27 of 2020 were randomly selected and entered in our study. Two sets of naso- and oropharyngeal swabs were collected, one set by a lab technician and the other by the patients, and the COVID-19 rRT-PCR test was performed. RESULTS: Of 50 selected cases, in seven patients all collected naso- and oropharyngeal swabs tested positive, and in 22 patients all samples tested negative for COVID-19 in rRT-PCR. Discrepancies between rRT-PCR results of lab technician- and patient-collected swabs were observed in 12 nasopharyngeal and 13 oropharyngeal specimens. Positive lab technician-collected and negative patient-collected samples were observed in 10 and 5 nasopharyngeal and oropharyngeal specimens, respectively. Negative lab technician-collected and positive patient-collected samples were observed in two and seven nasopharyngeal and oropharyngeal specimens, respectively. The overall percentage of agreement among both nasopharyngeal and oropharyngeal swabs taken by a lab technician and patients was 76% with a kappa value of 0.49 (P=0.001). CONCLUSION: Based on our findings, lab technician-collected naso- and oropharyngeal swabs cannot be replaced by patient-collected ones with regard to COVID-19 rRT-PCR.

Determining the association of thrombophilic gene polymorphisms with recurrent pregnancy loss in Iranian women.

OBJECTIVE: Thrombophilia is known to be associated with poor pregnancy outcomes. In this study, three thrombophilic gene polymorphisms, including EPCR (Ser219Gly), F11 (rs4253417) and F7 (323 Ins10) were investigated in an Iranian population of women in order to determine the correlation between thrombophilia and recurrent pregnancy loss (RPL). METHODS: Polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) were used to evaluate the frequency of three candidate thrombophilic risk factors for recurrent pregnancy loss. The frequencies of the polymorphisms were compared between the case (144 patients with a history of at least two miscarriages) and the control (150 healthy women with no abortion) group. RESULTS: Our results show that EPCR and FVII polymorphisms of the patient and control group have the same genotype frequency, and the difference is not statistically significant (p-value > .05). Regarding FXI polymorphism, TT genotype frequency was higher in the patient group than the control group (p-value < .05); however, CT heterozygote form was higher in the control group compared to the patient group (p-value < .05). CONCLUSION: In FXI polymorphism, T allele is possibly an RPL risk factor and C allele has a protective role. Thus, wild type FXI could be related to RPL, but EPCR and FVII polymorphism have no such correlation.

Residual methylation of tumor suppressor gene promoters, RASSF6 and RASSF10, as novel biomarkers for minimal residual disease detection in adult acute lymphoblastic leukemia.

Aberrant promoter methylation of RASSF6 and RASSF10 occurs at a high frequency in acute lymphoblastic leukemia (ALL). Because of the complexity of the current minimal residual disease (MRD) detecting-methods, the DNA methylation status of the RASSF6 and RASSF10 genes could potentially become biomarkers for the assessment of MRD levels in ALL patients. The promoter methylation status of RASSF6 and RASSF10 was assessed by using methylation-specific PCR (MSP) in the DNA isolated from 280 peripheral blood (PB) samples taken at the time of diagnosis, day 14, 28, and from the subsequent 30-month follow-ups of 45 adult ALL patients. The relative methylation level obtained during the follow-ups by MSP was compared to the MRD results obtained by the amplification of IG/TCR clonal rearrangements using the allele-specific quantitative-PCR (ASO-PCR) assay. Frequently, RASSF6 was methylated in B-ALL, and RASSF10 was methylated in T-ALL. The applicability and accuracy of the assays were increased when these markers were combined (91.1% and 93.8%, respectively). When a cutoff was defined for the PCR-MRD level, results of the 30 months of MRD detection showed a significant correlation between the PCR and MSP techniques (r = 0.848; p < 0.001). Due to the high applicability, the non-invasiveness, and promising prospect of longitudinal assessment, the DNA methylation status of the RASSF6 and RASSF10 genes could be potential biomarkers for the assessment of residual disease in PB of patients with ALL.

Effect of a high-intensity interval training on serum microRNA levels in women with breast cancer undergoing hormone therapy. A single-blind randomized trial.

BACKGROUND: The role of microRNAs (miRs) in hormone therapy (HT) is of keen interest in developing biomarkers and treatments for individuals with breast cancer. Although miRs are often moderate regulators under homeostatic conditions, their function is changed more in response to physical activity. OBJECTIVE: This single-blind randomized trial aimed to explore the effect of high-intensity interval training (HIIT) on serum levels of miRs in individuals with early-stage breast cancer undergoing HT. METHODS: Hormone receptor-positive women with breast cancer and healthy women were randomly assigned to a healthy control group (n=15), healthy group with HIIT (n=15), breast cancer group with HT (HT, n=26), and breast cancer group with HT and HIIT (HT+HIIT, n=26). The exercise groups underwent interval uphill walking training on a treadmill 3 times a week for 12weeks. At the end of the study, we analyzed changes in levels of cancer-related miRs (oncomiRs) and tumour suppressor miRs (TSmiRs) in response to the HT and HIIT. RESULTS: In women with breast cancer versus healthy controls, the expression of some oncomiRs was significantly increased - miR-21 (P<0.001), miR-155 (P=0.001), miR-221 (P=0.008), miR-27a (P<0.001), and miR-10b (P=0.007) - and that of some TSmiRs was significantly decreased - miR-206 (P=0.048), miR-145 (P=0.011), miR-143 (P=0.008), miR-9 (P=0.020), and let-7a (P=0.005). Moreover, HT considerably downregulated oncomiRs and upregulated TSmiRs. HIIT for 12weeks with HT significantly decreased the expression of the oncomiRs and significantly increased that of the TSmiRs as compared with HT alone. CONCLUSIONS: HITT could amplify the decrease and/or increase in expression of miRs associated with HT in women with breast cancer. A prospective trial could determine whether the use of circulating miRs for monitoring treatment can be useful in therapy decisions. TRIAL REGISTRATION: Iranian Registry of Clinical Trials (No.: IRCT201202289171N1).

Expression and CpG island methylation pattern of MMP-2 and MMP-9 genes in patients with congenital factor XIII deficiency and intracranial hemorrhage.

OBJECTIVES: Congenital factor XIII (FXIII) deficiency is a rare severe bleeding disorder. Intracranial hemorrhage (ICH) is the leading cause of mortality and morbidity in FXIII deficiency. However, its pathogenesis is not well understood yet. In this study, we investigated the expression and CpG island methylation status of matrix metalloproteinase-2 (MMP-2) and MMP-9 in patients with FXIII deficiency and ICH. METHODS: Forty patients with FXIII deficiency including twenty patients with ICH, and twenty without ICH were recruited as case and control groups, respectively. Methylation status was determined by bisulfite sequencing polymerase chain reaction (PCR), and gene expression was assessed by quantitative real-time PCR. RESULTS AND DISCUSSION: We found an unmethylated pattern for both MMP-2 and MMP-9 genes in the case group. Both genes were partially methylated in the control group, while the percentage of methylated CpGs was significantly higher in MMP-9 than MMP-2 (P = 0.001). Furthermore, higher expression of MMP-9 (in both the mRNA and protein levels) was found in the case than control group (P = 0.008 and P = 0.009, respectively). On the other hand, there was no significant difference in MMP-2 expression level (neither mRNA nor protein) between the two groups (P = 0.12 and P = 0.25, respectively). CONCLUSION: Our findings indicated that MMP-9 over-expression might be related to ICH in FXIII deficiency, and gene methylation effectively regulates its expression. Future researches will expand our understanding of the pathogenesis of ICH in congenital FXIII deficiency.

Generation of Induced Pluripotent Cancer Cells from Glioblastoma Multiform Cell Lines.

Generation of induced pluripotent stem cells (iPSCs) has been described as a powerful method to dedifferentiate the specialized cells to pluripotency. However, obtaining cancer-specific iPS cells (iPCs) encounters several barriers. The generation of iPCs provides valuable experimental platforms to mimic oncogenesis and offers potentials regarding drug screening. To overcome the difficulties regarding the iPC generation, we aimed at optimizing the generation of iPCs from glioblastoma multiform (GBM) cell lines and at understanding the potential barriers ahead of this process. The T731, T653, and mouse embryonic fibroblast cells were transduced by using retroviral plasmids encoding Oct4, Sox2, and Klf4. The cells were cultured on a layer of feeder cells for 14 days in iPS media and the obtained colonies were then picked and expanded to be evaluated for pluripotency markers by alkaline phosphatase staining, qRT-PCR, and Western blotting. Our findings confirmed resistance in cancer cells to achieve the pluripotency markers. In addition to designing technical tricks to obviate the barriers ahead of iPC generation, we suggested the small molecule PD98059 to enhance the efficiency of iPC generation from GBM cell lines. The resulting iPCs can further be used as a platform to study the mechanism of cancer formation and as a tool for drug screening for the treatment of patients with GBM.