Combination of Etoposide and quercetin-loaded solid lipid nanoparticles Potentiates apoptotic effects on MDA-MB-231 breast cancer cells.

Background: Breast cancer is a major global cancer, for which radiation and chemotherapy are the main treatments. Natural remedies are being studied to reduce the side effects. Etoposide (ETO), a chemo-drug, and quercetin (QC), a phytochemical, are considered potential factors for adaptation to conventional treatments. Objectives: The anticancer effect of the synergy between ETO and Quercetin-loaded solid lipid nanoparticles (QC-SLNs), was investigated in MDA-MB-231 cells. Methods: We developed QC-SLNs for efficient cellular delivery, characterizing their morphology, particle size, and zeta potential. We assessed the cytotoxicity of QC-SLNs and ETO on breast cancer cells via the MTT assay. Effects on apoptosis intensity in MDA-MB-231 cells have been detected utilizing annexin V-FITC, PI, and caspase activities. Real-time PCR assessed Bax gene and Bcl-2 gene fold change expression, while Western blot analysis determined p53 and p21 protein levels. Results: Spherical, negatively charged QC-SLNs, when combined with ETO, significantly enhanced inhibition of MDA-MB-231 cell proliferation compared to ETO or QC-SLNs alone. The combined treatment also notably increased the apoptosis pathway. QC-SLNs + ETO increased the Bax/Bcl-2 gene ratio, elevated p53 and p21 proteins, and activated caspase 3 and 9 enzymes. These results indicate the potential for QC-SLNs + ETO as a strategy for breast cancer treatment, potentially overcoming ETO-resistant breast cancer chemoresistance. Conclusion: These results suggest that QC-SLN has the potential to have a substantial impact on the breast cancer cure by improving the efficacy of ETO. This enhancement could potentially help overcome chemoresistance observed in ETO-resistant breast cancer.

Comparison of biosynthetic zinc oxide nanoparticle and glucantime cytotoxic effects on Leishmania major (MRHO/IR/75/ER).

Currently, zinc oxide (ZnO) particles are used in nanotechnology to destroy a wide range of microorganisms. Although pentavalent antimony compounds are used as antileishmanial drugs, they are associated with several limitations and side effects. Therefore, it is always desirable to try to find new and effective treatments. The aim of this research is to determine the antileishmanial effect of ZnO particles in comparison to the Antimoan Meglumine compound on promastigotes and amastigotes of Leishmania major (MRHO/IR/75/ER). After the extraction and purification of macrophages from the peritoneal cavity of C57BL/6 mice, L. major parasites were cultured in Roswell Park Memorial Institute-1640 culture medium containing fetal bovine serum (FBS) 10% and antibiotic. In this experimental study, the effect of different concentrations of nanoparticles was investigated using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) colorimetric method, in comparison to the glucantime on promastigotes, amastigotes and healthy macrophages in the culture medium. The amount of light absorption of the obtained color from the regeneration of tetrazolium salt to the product color of formazan by the parasite was measured by an enzyme-linked immunosorbent assay (ELISA) reader, and the IC50 value was calculated. IC50 after 24 h of incubation was calculated as IC50 = 358.6 µg/mL. The results showed, that the efficacy of ZnO nanoparticles was favorable and dose-dependent. The concentration of 500 µg/mL of ZnO nanoparticles induced 84.67% apoptosis after 72. Also, the toxicity of nanoparticles was less than the drug. Nanoparticles exert their cytotoxic effects by inducing apoptosis. They can be suitable candidates in the pharmaceutical industry in the future.

Methylation-mediated silencing of miR-125a-5p facilitates breast cancer progression by inducing autophagy.

BACKGROUND: microRNA-125a-5p (miR-125a) is a tumor suppressor gene whose role in autophagy remains poorly understood. In the current study, we aimed to investigate the methylation status of miR-125a, its transfection into SK-BR3 cells, and its effects on autophagy. METHODS: Sixty samples of tumor and non-tumor adjacent tissue were collected and the methylation status of miR-125a was evaluated by methylation-specific PCR (MSP). The effect of 5-Aza-dC on miR-125a expression was investigated in the SK-BR3 cells. Cells were also transfected with miR-125a mimic/antimiR. The expression of miR-125a and its target genes was evaluated by Real-Time PCR. Protein levels of ATG5 and LC3 were assessed by Western blotting. HER2 expression was investigated by immunocytochemistry (ICC). RESULTS: The data showed that the miR-125a promoter CpG Island was significantly hypermethylated in breast cancer tissues (p < 0.01) and in SK-BR3 cells. The 5-Aza-dC could significantly increase miR-125a expression by decreasing its methylation (p < 0.05). In addition, Western blot analysis indicated the expression of ATG5 and LC3 II/ LC3I, as autophagy biomarkers, was significantly reduced in SK-BR3 cells transfected with miR-125a (p < 0.05). CONCLUSIONS: Our data showed miR-125a expression was significantly decreased in tumor tissues due to its promoter hypermethylation. Overexpression of miR-125a was associated with a reduction in autophagy, which could provide a new therapeutic avenue for advanced-stage breast cancer treatment.

Molecular and Serological Prevalence of HCMV in Iranian Patients with Breast Cancer.

BACKGROUND: Human cytomegalovirus (HCMV) is prevalent viral infection involved in several human cancers including breast cancer. The presence of HCMV genome in breast cancer tissue and footprint of viral last exposure patient's serum are considered as important factor in the process of breast cancer development. OBJECTIVES: This study aimed to investigate molecular and serological epidemiology of HCMV in patients with breast cancer in Iran for first time. METHODS: In our case-control study, 98 samples of breast tissue, including 49 cancerous (case) and 49 adjacent non-cancerous tissue were collected (control). In addition, we collected sera samples from all patients (n=49) and healthy individual (n=49). Seroprevalence of HCMV was assessed by Enzyme-linked immunosorbent assay (ELISA) and detection of HCMV genome was performed using Nested-PCR method. RESULTS: HCMV genome found in 16.3% (8/49) of cases tissue and 2% (1/49) of controls tissue. In patients group, the levels of anti-CMV IgG and IgM were 93.9% and 2% compared to 69.4% and 4.1% in healthy individuals, respectively. There was a statistically difference between the anti-CMV IgG in patients and healthy control (p= 0.002). We found 75% of (6/8) HCMV genome positive PCR samples were also positive for their anti-CMV IgG in cases which was statistically significant (p= 0.01).  Conclusions: Our result showed significant presence of HCMV genome and anti-CMV IgG in patients, supporting the role of HCMV in breast cancer.
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Conditional disease-free survival rates and their associated determinants in patients with epithelial ovarian cancer: A 15-year retrospective cohort study.

BACKGROUND: The most common type of ovarian cancer (OC) is epithelial ovarian cancer (EOC) which is the most lethal gynecologic malignancy in adult women. AIM: This study aimed to determine the conditional disease-free survival (CDFS) rates and their associated determinants in patients with EOC. METHODS AND RESULTS: The clinical and demographic data of 335 patients with confirmed EOC at Motahari Clinic (Shiraz, Iran) were retrospectively reviewed and analyzed. Traditional DFS (TDFS) and CDFS were calculated using the Kaplan-Meier method and cumulative DFS estimates, respectively. To evaluate the effects of the prognostic determinants on the DFS of the patients, a multiple covariate Cox analysis using the landmarking method was applied. The 1- and 3-year TDFSs were 81.1% and 47.0%, respectively, and decreased over time. At baseline, a higher stage tumor and endometrioid histology were associated with a higher risk of recurrence when compared to stage I and other histological subtypes, respectively. The hazard of recurrence for older women (age ≥55 years) was approximately twice and three times more than that of women aged <45 years at 1- and 3-year landmark time points, respectively. CONCLUSION: The age at diagnosis, defined by a cut-off of 55 years, was a prognostic factor for the CDFS of EOC women. Moreover, patients with advanced-stage EOC (ASEOC) (stages III and IV) and endometrioid histology had poorer CDFSs compared to those with early-stage EOC (ESEOC) (stages I and II) and other histological types. In ESEOC patients with age at diagnosis of >55 years, CDFS gradually decreased in 3 years after remission which should be considered for follow-up care decision-making.

Evaluation of microRNA-223 and microRNA-125a expression association with STAT3 and Bcl2 genes in blood leukocytes of CLL patients: a case-control study.

OBJECTIVE: In chronic lymphocytic leukemia (CLL), lack of expression or dysregulation of some special miRs disrupts apoptosis of malignant cells; thereby miR expression can enhance cell proliferation, disease progression and decrease patient survival. RESULTS: 30 CLL patients and 20 healthy individuals participated in the study. RNA was extracted to evaluate the expression of miR-125, miR-223, BCL-2 and signal transducer and transcription 3 activator (STAT3) genes; quantitative Real Time- PCR (Q-RT-PCR) was performed. MiR-125a and miR-223 expression decreased in the patients compared to the control group (P-Value:0.001). BCL-2 and STAT3 which are the target genes of these two miRs, showed increased expression, in the patients compared to the control subjects (P-Value: 0.001 and P-Value: 0.64 respectively). A significant reverse relationship was found between miR-125a and BCl-2 expression and WBC count. Significantly, miR-223 expression was associated with smoking in patients (P-Value: 0.007). Also, these miRs may have regulatory effects by controlling white blood cell (WBC) production based on the inverse correlation with WBC count and hemoglobin (Hb) concentration. Finally, miR-223 can be used as a prognostic factor in CLL patients; miR-125a may be useful for evaluating the therapeutic approaches based on the inverse link with BCl-2.

Identical Twins with a Mutation in the STK4 Gene Showing Clinical Manifestations of the Mutation at Different Ages: A Case Report.

Combined immunodeficiencies (CIDs) are a heterogeneous group of disorders characterized by various gene mutations. The mutations in the STK4 (Serine Threonine Kinase 4) gene, which has a role in the regulation of apoptosis and proliferation, can be a cause of immunodeficiency. In the current paper, we reported a case of identical twin brothers with a novel STK4 mutation, one of whom showed clinical manifestations associated with this mutation with a delay of two years. The mutation in the STK4 gene was identified employing Whole Exome Sequencing (WES), and we described the probable reasons for this delay. We found that the STK4 genetic defect caused almost the same clinical symptoms of immunodeficiency in the twin brothers. Meanwhile, the severity of the disease was higher in one of them, which may be due to extra genetic defect in LRBA, and likely differences in the percentage of B lymphocyte population and CD4+/ CD8+ state.

Tumor-Induced Metabolism and T Cells Located in Tumor Environment.

Several subtypes of T cells are located in a tumor environment, each of which supplies their energy using different metabolic mechanisms. Since the cancer cells require high levels of glucose, the conditions of food poverty in the tumor environment can cause inactivation of immune cells, especially the T-effector cells, due to the need for glucose in the early stages of these cells activity. Different signaling pathways, such as PI3K-AKt-mTOR, MAPK, HIF-1α, etc., are activated or inactivated by the amount and type of energy source or oxygen levels that determine the fate of T cells in a cancerous environment. This review describes the metabolites in the tumor environment and their effects on the function of T cells. It also explains the signaling pathway of T cells in the tumor and normal conditions, due to the level of access to available metabolites and subtypes of T cells in the tumor environment.

The up-regulation of markers of adipose tissue fibrosis by visfatin in pre-adipocytes as well as obese children and adolescents.

Adipocytes are surrounded by a three-dimensional network of extracellular matrix (ECM) proteins. Aberrant ECM accumulation and remodeling leads to adipose tissue fibrosis. Visfatin is one of the adipocytokines that is increased in obesity and is implicated in insulin resistance. The objective of this study was to investigate the effect of visfatin on major components of ECM remodeling. In this study, plasma levels of both endotrophin and visfatin in obese children and adolescents were significantly higher than those in control subjects and they showed a positive correlation with each other. Treatment of 3T3-L1 pre-adipocytes with visfatin caused significant up-regulation of Osteopontin (Opn), Collagen type VI (Col6), matrix metalloproteinases MMP-2 and MMP-9. By using inhibitors of major signaling pathways it was shown that visfatin exerted its effect on Col6a3 gene expression through PI3K, JNK, and NF-кB pathways, while induced Opn gene expression via PI3K, JNK, MAPK/ERK, and NOTCH1. Our conclusion is that, the relationship between visfatin, endotrophin and insulin resistance parameters in obesity as well as increased expression of ECM proteins by visfatin suggests adipose tissue fibrosis as a mechanism for devastating effects of visfatin in obesity.

Parity as a Prognostic Factor in Patients with Advanced-Stage Epithelial Ovarian Cancer.

AIM: This study aimed to determine the prognostic factors influencing the overall survival (OS) of Iranian women with epithelial ovarian cancer (EOC). METHODS: Information about newly diagnosed patients with confirmed EOC at Motahari Clinic, Shiraz, Iran, from January 1, 2001, to December 31, 2016, was retrospectively reviewed and analyzed. Cox-adjusted proportional hazards (PH) and stratified Cox (SC) models were used to determine the potential prognostic factors. RESULTS: The mean (±SD) age at the diagnosis of 385 patients with EOC was 49.0 (±13.2) years old. Early-stage EOC (ESEOC) and advanced-stage EOC (ASEOC) were diagnosed in 34.3% and 65.7% of the total patients, respectively. The median (95% CI) OS was 35 (28-41) months. For ESEOC patients, a stage II-tumor led to a lower OS in the multivariable analysis compared to a lower stage tumor (P= 0.025). For ASEOC patients, age≥65 years at diagnosis (P=0.008) led to a lower OS. ASEOC patients with 2-5 parities (P=0.014) and >5 parity (P=0.001) demonstrated better OS than nulliparous women. CONCLUSION: Patients with ESEOC, higher tumor stage was associated with a shorter OS. The age at diagnosis harmed the OS of patients with ASEOC. More than one parity improved OS in ASEOC patients.

The potential role of miR-29 in health and cancer diagnosis, prognosis, and therapy.

miR-29 family is one of the small noncoding RNAs and has a very important role in many physiologic and pathologic functions through regulating the target genes that play roles in various bioprocesses such as proliferation, survival, apoptosis, and angiogenesis. Thus, we aim to survey the potential of the miR-29 family in normal model and development and progression of malignancy in this study. In addition, the potential role of miR-29 family has been studied as the clinical marker for the diagnosis and prognosis of many cancers as the potential targets to treat cancer. Moreover, it was stated in summary that the herbal compounds can regulate miR-29 family in cancers. Therefore, regulating the expression of the miR-29 family in a variety of cancers can be a new strategy to obtain better results from cancerous patients' treatment in the future.

Evaluation of Plasma TRB3 and Sestrin 2 Levels in Obese and Normal-Weight Children.

OBJECTIVE: Obesity in childhood and adolescence is associated with metabolic syndrome and cardiovascular diseases. TRB3 (Tribbles homolog 3) and sestrin 2 are two newly found proteins that have been identified to play an important role in obesity and its complications. AIM: The purpose of this study was to evaluate concentrations of TRB3 and sestrin 2 in plasma of obese and normal-weight children and adolescents, and their association with metabolic and anthropometric parameters. METHODS: Plasma levels of TRB3, sestrin 2, insulin, fasting plasma glucose (FPG), and lipid profile were evaluated in 70 children and adolescents (34 obese and 36 controls). Insulin resistance was calculated using a homeostasis model assessment of insulin resistance. Metabolic syndrome was defined according to IDF criteria. RESULTS: Plasma TRB3 levels of the obese subjects were significantly higher than that of normal weight subjects. TRB3 levels were positively correlated with BMI, BMI z-score, waist circumference, and FPG. The concentration of sestrin 2 was significantly lower in obese subjects compared to normal-weight subjects. A statistically significant positive correlation was observed between plasma concentrations of sestrin 2 and high-density lipoprotein cholesterol. Neither TRB3 nor sestrin 2 were correlated with insulin resistance and metabolic syndrome. CONCLUSION: Both TRB3 and sestrin 2 may contribute to the development of obesity and its complications and can be considered interesting therapeutic target for the treatment of obesity.

Two novel mutations in the KHDC3L gene in Asian patients with recurrent hydatidiform mole.

Recurrent hydatidiform mole (RHM) is defined by the occurrence of repeated molar pregnancies in affected women. Two genes, NLRP7 and KHDC3L, play a causal role in RHM and are responsible for 48-80% and 5% of cases, respectively. Here, we report the results of screening these two genes for mutations in one Iranian and one Indian patient with RHM. No mutations in NLRP7 were identified in the two patients. KHDC3L sequencing identified two novel protein-truncating mutations in a homozygous state, a 4-bp deletion, c.17_20delGGTT (p.Arg6Leufs*7), in the Iranian patient and a splice mutation, c.349+1G>A, that affects the invariant donor site at the junction of exon 2 and intron 2 in the Indian patient. To date, only four mutations in KHDC3L have been reported. The identification of two additional mutations provides further evidence for the important role of KHDC3L in the pathophysiology of RHM and increases the diversity of mutations described in Asian populations.

The genomic architecture of NLRP7 is Alu rich and predisposes to disease-associated large deletions.

NLRP7 is a major gene responsible for recurrent hydatidiform moles. Here, we report 11 novel NLRP7 protein truncating variants, of which five deletions of more than 1-kb. We analyzed the transcriptional consequences of four variants. We demonstrate that one large homozygous deletion removes NLRP7 transcription start site and results in the complete absence of its transcripts in a patient in good health besides her reproductive problem. This observation strengthens existing data on the requirement of NLRP7 only for female reproduction. We show that two other variants affecting the splice acceptor of exon 6 lead to its in-frame skipping while another variant affecting the splice donor site of exon 9 leads to an in-frame insertion of 54 amino acids. Our characterization of the deletion breakpoints demonstrated that most of the breakpoints occurred within Alu repeats and the deletions were most likely mediated by microhomology events. Our data define a hotspot of Alu instability and deletions in intron 5 with six different breakpoints and rearrangements. Analysis of NLRP7 genomic sequences for repetitive elements demonstrated that Alu repeats represent 48% of its intronic sequences and these repeats seem to have been inserted into the common NLRP2/7 primate ancestor before its duplication into two genes.

Selenium and its relationship with selenoprotein P and glutathione peroxidase in children and adolescents with Hashimoto's thyroiditis and hypothyroidism.

The essential trace element selenium (Se) is required for thyroid hormone synthesis and metabolism. Selenoproteins contain selenocysteine and are responsible for biological functions of selenium. Glutathione peroxidase (GPx) is one of the major selenoproteins which protects the thyroid cells from oxidative damage. Selenoprotein P (SePP) is considered as the plasma selenium transporter to tissues. The aim of this study was to evaluate serum Se and SePP levels, and GPx activity in erythrocytes of children and adolescents with treated Hashimoto's thyroiditis, hypothyroidism, and normal subjects. Blood samples were collected from 32 patients with Hashimoto's thyroiditis, 20 with hypothyroidism, and 25 matched normal subjects. All the patients were under treatment with levothyroxine and at the time of analysis all of the thyroid function tests were normal. GPx enzyme activity was measured by spectrophotometry at 340 nm. Serum selenium levels were measured by high-resolution continuum source graphite furnace atomic absorption. SePP, TPOAb (anti-thyroid peroxidase antibody), and TgAb (anti-thyroglobulin antibody) were determined by ELISA kits. T4, T3, T3 uptake and TSH were also measured. Neither GPx activity nor SePP levels were significantly different in patients with Hashimoto's thyroiditis or hypothyroidism compared to normal subjects. Although GPx and SePP were both lower in patients with hypothyroidism compared to those with Hashimoto's thyroiditis and normal subjects but the difference was not significant. Serum Se levels also did not differ significantly in patients and normal subjects. We did not find any correlation between GPx or SePP with TPOAb or TgAb but SePP was significantly correlated with Se. Results show that in patients with Hashimoto's thyroiditis or hypothyroidism who have been under treatment with levothyroxine and have normal thyroid function tests, the GPx, SePP and Se levels are not significantly different.

Prevalence of liver fluke infections in slaughtered animals in Lorestan, Iran.

Fasciola spp. and Dicrocoelium dendriticum as liver flukes, contaminate ruminants and other mammalian extensively and cause major diseases of livestock that create considerable economic losses. This retrospective study has been done to evaluate contamination rate of slaughtered animals with fasciolosis and dicrocoeliosis at Lorestan abattoirs. In this survey, prevalence rate of fasciolosis and dicrocoeliosis in slaughtered animals in a 3-year period (2010-2013) has been analyzed. A total of 356,605 livestock including 265,692 sheep and 90,913 goats were slaughtered in the 3-year period and overall 39,613 (11.1 %) livers were condemned. Fascioliasis and dicrocoeliosis were responsible for 6.3 and 4.8 % of total liver condemnations in this period, respectively. Fasciola spp. and D. dendriticum infection in sheep (7.1 and 5.6 %, respectively) were considerably higher than goats (3.9 and 2.6 %, respectively). The annual prevalence rates showed a significant decline in the fasciolosis and dicrocoeliosis infection in goats (p < 0.001). Data showed significant seasonal pattern for distomatosis in sheep and goats (p < 0.001). Liver condemnations due to fasciolosis were prevalent in sheep and goats slaughtered during spring and autumn, respectively, whereas dicrocoeliosis were common in spring season for both sheep and goats. This survey provides baseline data for the future monitoring of these potentially important parasitic infections in the region.

Evaluation of copper bioaccumulation and translocation in Jatropha curcas grown in a contaminated soil.

Contamination of soils with copper (Cu) has become a serious problem in the environment. Phytoremediation is an emerging green technology that uses green plants to remediate heavy metal contaminated areas. This study was conducted to evaluate the potential of Jatropha curcasfor remediation of soils contaminated with Cu. Seedlings were planted in soils spiked with Cu in amount of 0, 50, 100, 200, 300, and 400 mg kg(-1) (Cu0, Cu50, Cu100, Cu200, Cu300, and Cu400) for a period of five months. The maximum height and number of leaves were recorded in control (Cu0) whereas the highest basal stem diameter was found in seedlings exposed to Cu50. Copper concentrations among plant parts were in the following trend: roots > stems > leaves. The highest total Cu concentration (665 +/- 1 mg kg(-1)) and total Cu removal (1.2 +/- 0.2%) based on total plant dry biomass were found in Cu400 and Cu50 treatments, respectively. J. curcas exhibited high root concentration factor (RCF > 1) and low translocation factor (TF < 1). Although Cu accumulation by the plant didn't reach the criteria of Cu hyperaccumulators, this species showed a potential to be used in phytostabilization of mildly Cu contaminated areas. However, the plant cannot be used for phytoextraction of Cu-contaminated soils.