Evaluation of antioxidant status and oxidative stress markers in HTLV-1 infected individuals: correlation with the severity of virus-induced complications.

Introduction. Human T-cell lymphotropic virus type 1 (HTLV-1), a well-known member of the retroviridae family, potentially causes serious outcomes including adult T-cell leukaemia/lymphoma (ATLL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM-TSP). Oxidative stress plays a key role in progression and clinical exacerbation of several chronic infections. We have previously shown a reduction in serum total antioxidant capacity (TAC) during HTLV-1 infection and this study was set out to investigate the reasons for TAC reduction.Hypothesis/Gap Statement. Oxidant/antioxidant imbalance during HTLV-1 infection may result from disruptions in oxidant levels or antioxidant defence system.Aim. This study aimed to analyse the key enzymes and oxidant molecules playing important roles in virus-induced oxidative stress.Methodology. We measured serum activities of the major antioxidant enzymes; superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX) as well as serum concentrations of the main oxidant markers: nitric oxide (NO) and malondialdehyde (MDA). Totally 40 HTLV-1 infected patients and 40 healthy controls were enrolled in this study. The patient group consisted of chronic carriers and patients with HAM-TSP (N=20).Results. The current study found that serum levels of MDA and NO were significantly higher in patient groups particularly in HAM-TSP patients (P<0.05). In addition, a reductive trend was observed in the serum activities of CAT, SOD, and GPX in HTLV-1 infected patients compared with healthy controls (P<0.05).Conclusion. Reduced activities of CAT, SOD, and GPX antioxidant enzymes along with the observed elevated concentrations of oxidant molecules may contribute to oxidative stress and worse outcomes during HTLV-1 infection.

Aprepitant Promotes Caspase-Dependent Apoptotic Cell Death and G2/M Arrest through PI3K/Akt/NF-κB Axis in Cancer Stem-Like Esophageal Squamous Cell Carcinoma Spheres.

The antagonists of the neurokinin-1 receptor (NK1R) are known for their anti-inflammatory, anxiolytic, antiemetic, and anticancer activities. Aprepitant, a nonpeptide NK1R antagonist, is used in nausea and vomiting, the most common side effects of cancer chemotherapy in patients. It has been established that NK1R activation by substance P (SP), which links cancer promotion and progression to a neurokinin-mediated environment, became one mechanism that corresponds to the mitogenesis of tumor cells. Therefore, this study is aimed at explaining and evaluating the anticancer impacts of aprepitant on esophageal squamous cancer cell (ESCC) spheres by using in vitro experiments, such as resazurin, ROS, annexin-V binding, RT-PCR, and Western blot analysis. As a result, we showed that aprepitant had strong antiproliferative and cytotoxic effects on ESCC cell spheres. Also, aprepitant caused significant G2-M cell cycle arrest depending on concentration increase. Further, exposure of cells to this agent resulted in caspase -8/-9-dependent apoptotic pathway activation by modifying the expression of genes involved in apoptosis. Besides, treatment of the cells by aprepitant abrogates of the PI3K/Akt pathway, as shown by reducing the level of Akt, induces apoptotic cell death. In summary, pharmacological inhibition of NK1R with aprepitant seems to have a significant chance of treating ESCC as a single agent or in conjunction with other chemotherapeutic drugs.

Higher detection of JC polyomavirus in colorectal cancerous tissue after pretreatment with topoisomerase I enzyme; colorectal tissue serves as a JCPyV persistence site.

BACKGROUND: The JC polyomavirus has been blamed to contribute in colorectal cancer (CRC), however, the topic is still controversial. Varying detection rate of JCPyV genome has been reported mainly due to technical reasons. Here, we provide summative data on the topic, with emphasize on technical issues. METHODS: Formalin-fixed paraffin-embedded tissue samples from 50 patients with CRC, consisting of tumoral and non-cancerous marginal tissue (totally 100 samples) were included in the study. After DNA extraction, specific JCPyV T-Ag sequences were targeted using Real-time PCR. To unwind the supercoiled JCPyV genome, pretreatment with topoisomerase I, was applied. Immunohistochemical (IHC) staining was performed using an anti-T-Ag monoclonal antibody. RESULTS: In the first attempts, no samples were found to be positive in Real-time PCR assays. However, JCPyV sequences were found in 60% of CRC tissues and 38% of non-cancerous colorectal mucosa after application of pre-treatment step with topoisomerase I enzyme (P = 0.028). T-Ag protein was found in the nuclear compartment of the stained cells in IHC assays. CONCLUSIONS: The presence of JCPyV in CRC tissues, as well as T-Ag localization in the nucleolus, where its oncogenic effect takes place, may provide supporting evidence for JCPyV involvement in CRC development. The study highlights the importance of using topoisomerase I to enhance JCPyV genome detection. Also, colorectal tissue is one of the permissive human tissue for JC resistance after preliminary infection.

Evaluation of glutathione reductase activity in colon tissue of patients with irritable bowel syndrome.

OBJECTIVES: Irritable bowel syndrome (IBS) is known as one of the most common irritating gastrointestinal disorders. The mechanism behind IBS is still under investigation and it is thought that it may arose from multi factors among which free radicals have been previously mentioned. Studies have found an association between oxidative stress and IBS; however, little is known about the mechanisms and oxidative stress components status during IBS. One of the key factors playing a central role in oxidative stress network is glutathione reductase (GR). Here we report the GR activity in colon tissue samples during IBS to explore a part of contributing components in IBS pathogenesis. METHODS: The GR enzyme activity was measured in 15 active IBS colon biopsy samples and was compared to our best available age and sex matched colorectal tissue samples from normal marginal tissue of resected colon cancers (n=15). The enzyme activity in the two groups was determined and compared using a commercial GR Assay Kit (Cayman chemical). RESULTS: A significant decrease in GR activity among IBS tissue samples was observed compared to anatomically normal marginal colon tissue samples (p=0.007). CONCLUSIONS: Lower GR activity may increase oxidized glutathione there by in turn could contribute as a main component in oxidative stress network. The lower GR activity results in hampered defense mechanism against produced free radical species. This finding may clarify a part of IBS pathogenesis.

MicroRNAs as potential investigative and predictive biomarkers in colorectal cancer.

Colorectal cancer (CRC) is a noticeable reason of cancer-associated deaths with a high incidence and mortality rate. Countless effort have been put into the improving clinical management of CRC patients including more effective tools and a wide variety of biomarkers for diagnostic, prognostic or predictive purposes. In recent years, dysregulated miRNAs have been emerged as highly sensitive and specific markers to manage CRC in an effective way. They can play key roles in carcinogenesis as potential oncogenes, tumor suppressors or regulators of cancer network. Therefore, miRNAs may serve as molecular tools that can be quantified and used in diagnostic and prognostic approaches. Growing evidence also suggests that forced expression of tumor suppressor miRNAs or inhibiting the oncogene ones, can be used as a novel treatment strategy. In this review, we focus on the clinical applications of miRNAs as promising biomarkers of early cancer detection, prognosis and treatment.

The role of substance P/neurokinin 1 receptor in the pathogenesis of esophageal squamous cell carcinoma through constitutively active PI3K/Akt/NF-κB signal transduction pathways.

One of the most prevalent malignancies is esophageal squamous cell carcinoma (ESCC), which is associated with high morbidity and mortality. Substance P (SP), as one of the peptides released from sensory nerves, causes the enhancement of cellular excitability through the activation of the neurokinin-1 (NK1) receptor in several human tumor cells. Aprepitant, a specific, potent, and long-acting NK1 receptor antagonist, is considered as a novel agent to inhibit proliferation and induce apoptosis in malignant cells. Since the antitumor mechanism of aprepitant in ESCC is not completely understood, we conducted this study and found that aprepitant induced growth inhibition of KYSE-30 cells and arrested cells in the G2/M phase of the cell cycle. Aprepitant also caused apoptotic cell death and inhibited activation of the PI3K/Akt axis and its downstream effectors, including NF-κB in KYSE-30 cells. Besides, quantitative real-time (qRT)-PCR analysis showed a significant down-regulation of NF-κB target genes in KYSE-30 cells, indicating a probable NF-κB-dependent mechanism involved in aprepitant cytotoxicity. Thus, the present study recommends that SP/NK1R system might, therefore, be considered as an emerging and promising therapeutic strategy against ESCC.

Intraperitoneal Administration of Telmisartan Prevents Postsurgical Adhesion Band Formation.

BACKGROUND: Angiotensin II receptor blockers (ARBs) have a potential role in reducing inflammation and fibrosis. We have integrated systems and molecular biology approaches to investigate the therapeutic potential of ARBs in preventing postsurgical adhesion band formation. MATERIAL AND METHODS: we have followed the ARRIVE guidelines point by point during experimental studies. Telmisartan (1 and 9 mg/kg), valsartan (1 and 9 mg/kg), and losartan (1 and 10 mg/kg) were administered intraperitoneally in different groups of male albino Wistar rat. After 7 d of treatment, macroscopic evidence and score of fibrotic bands based on scaling methods was performed. Moreover, the anti-inflammatory and antifibrosis effects of telmisartan on reduction of fibrotic bands were investigated by using histopathology, ELISA, and real-time polymerase chain reaction methods. RESULTS: Telmisartan, but not losartan or valsartan, prevented the frequency as well as the stability of adhesion bands. Telmisartan appears to elicit anti-inflammatory responses by attenuating submucosal edema, suppressing proinflammatory cytokines, decreasing proinflammatory cell infiltration, and inhibiting oxidative stress at the site of peritoneal surgery. We also showed that telmisartan prevents fibrotic adhesion band formation by reducing excessive collagen deposition and suppression of profibrotic genes expression at the peritoneum adhesion tissues. CONCLUSIONS: These results support the potential application of telmisartan in preventing postsurgical adhesion band formation by inhibiting key pathologic responses of inflammation and fibrosis in postsurgery patients.

The effect of curcumin in prevention of contrast nephropathy following coronary angiography or angioplasty in CKD patients.

INTRODUCTION: Contrast-induced nephropathy (CIN) is the most common cause of iatrogenic acute kidney injury. It is happened more commonly in patients with underlying kidney diseases. It is appeared that the oxidative stress is the main mechanism of contrast nephropathy. Curcumin is suggested as an herbal antioxidant agent, so we decided to assess the effect of curcumin in preventing of this complication in patients with underlying chronic kidney disease (CKD) who need coronary angiography. METHODS AND MATERIALS: We conducted double blind, placebo-controlled clinical trial in 60 moderate to severe CKD patients who underwent coronary angiography or angioplasty. Adjusted dose of Iodixanol was used as contrast agent in all of them. Curcumin or placebo administered orally, 1.5 g daily from 2 days before procedure to 3 days after it. CIN was defined by an increased serum creatinine level≥0.3mg/dl or an increase to ≥1.5 times of the baseline within 48 hours after procedure. Urinary NGAL test was also done the next day after angiography. RESULTS: CIN occurred in 12(20%) of patients, 5(16.7%) in Curcumin group and 7(23.3%) in placebo group (odds ratio [OR], 0.56; 95% confidence interval [CI], 0.18 to 2.36; P0.51). Serum Creatinine was increased after72 hours of intervention from 1.65±0.26 mg/dl to 1.79±0.33 mg/dl in Curcumin group and from 1.61±0.23 mg/dl to 1.86±0.35 in placebo group. There is no significant difference between the mean increase in serum creatinine concentration in the placebo group and Curcumin group (difference of 0.006 mg/dL; 95% CI, - 0.06 to 0.08; P0.85). Urinary NGAL test was significantly higher in patients with AKI (p=0.000), but there weren't differences in its level in two groups (p=0.761)  Conclusion: It is appeared prophylactic oral Curcumin hasn't protective effects on CIN in high risk patients who have undergone coronary procedure.

Molecular targeting for treatment of human T-lymphotropic virus type 1 infection.

Human T-cell lymphotropic virus type 1 (HTLV-1) infection is linked to adult T-cell leukemia-lymphoma (ATLL) and HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and several other disorders. ATLL occurs in approximately 5% of the 15-20 million people infected by HTLV-1 in the world. In general, ATLL is resistant to chemotherapy, which underlines the need for new and effective therapeutic strategies. Previous studies highlighted the role of viral enzymes, responsible for viral replication, and regulatory proteins such as Tax and HBZ in the progression of HTLV-1-associated diseases. There are conflicting reports on the efficacy of current enzyme inhibitors, mainly developed against human immunodeficiency virus (HIV), for treatment of HTLV-1 infection. New treatment approaches including monoclonal antibodies show promising results and exert significant cytotoxic effects on ATLL cells. This manuscript reviews the recent developments in molecular targeting for treatment of HTLV-1 infection.

The Status of Nitric Oxide and its Backup, Heme Oxygenase 1, in Thromboangiitis Obliterans.

BACKGROUND: Until recently, a gene polymorphism in the promoter region of endothelial nitric oxide synthase has been suggested as a risk factor for thromboangiitis obliterans (TAO) development. The aim of this study was to compare the metabolites of nitric oxide (NO) and its backup, heme-oxygenase-1 (HMOX1), between TAO patients and those of a smoking control group matched by race, age, sex, and smoking habits. METHODS: Twenty-four male Caucasian TAO patients and 20 male Caucasian controls enrolled in the study. Their smoking habits were matched based on the serum cotinine levels of 17 of the TAO patients and the 20 controls. A colorimetric kit was used to measure NO, and an enzyme-linked immunosorbent assay kit was used to measure cotinine and HMOX1 levels. RESULTS: The mean serum level of NO metabolites in the TAO group was significantly less than in the controls (p = 0.03) and also significantly less in the patients with below-knee amputations than in non-amputees (p= 0.018). Also, HMOX1 was significantly greater in the TAO patients than in the controls (p= 0.01). No significant correlation was found between NO and HMOX1 (p = 0.054). CONCLUSION: Nitric oxide may play a pivotal role in TAO development and its outcome. However, the intact HMOX1 pathway may demonstrate the unique role of NO, which cannot be compensated for by HMOX1 and whose absence may make patients susceptible to developing TAO. In addition, another pathway besides NO, with influence on vascular tone and hemostasis, might be involved in TAO development, such as the autonomic nervous system. Further studies are suggested regarding these issues.

Tissue Expression of Prohibition-I and It's Relationship With Prognostic Factors in Breast Cancer.

BACKGROUND & OBJECTIVE: Globally, breast cancer is the most common malignancy among females. Prohibition (PHB)-I, a homologous protein, was initially introduced as a suppressor gene for amplification process. Further, the protein has a key role in the cell cycle and is capable of inhibiting DNA transcription in many cell types. Therefore, its possible role in different types of human malignancies is of interest.The current study aimed at examining the relationship between the tissue distribution of PHB-I and prognostic factors of breast cancer. METHODS: Paraffin-embedded tissue specimens of 33 patients diagnosed with breast cancer at Omid teaching Hospital, Mashhad, Iran were studied and a commercial monoclonal antibody was used to perform immunohistochemistry (IHC). The relationship between PHB-I tissue expression with age, disease stage, tumor grade and size, as well as hormone receptor status including estrogen (ER) and progesterone (PR) receptors, and Her-2 receptor were evaluated. RESULTS: The Immunohistochemical analysis showed a relative increase in PHB-I tissue expression along with higher tumor grade (P=0.057). In addition, higher expression of ER and PR were observed (P=0.027 and 0.009, respectively). The age of patients and other prognostic factors including Her-2 receptor status and disease stage did not statistically correlate with PHB-I expression. CONCLUSION: An increased expression of PHB-I was observed in the breast cancer tumors of the current study patients compared with the anatomically healthy margin. Its coloration with some prognostic factors such as disease grade and expression of ER and PR might indicate the PHB-I potential application for diagnostic and patient management purposes.

Status of soluble ST2 levels in serum of HTLV-1 infected individuals.

ST2 is a member of IL-1 receptor family expressed on Th2 cells and regulates Th2 responces. The gene of ST2 encodes soluble ST2 (sST2) and the transmembrane ST2 (ST2L) isoforms through alternative mRNA splicing. The discovery of IL33/ ST2 signaling pathway, has drawn a great scientific attention to this system. sST2 has been shown to be an indacating factor in various infl ammatory conditions. This study aims to evaluate serum sST2 levels in HTLV-1 infected patients. This study included 49 HTLV-1 seropositive cases of which 14 were sympthomatic. Controls consisted of 30 healthy volunteers. sST2 level was measured using a quantitative ELISA assay and the results of the study groups were compared. Corroborating the previous reports, sST2 was lower in females (P = 0.003). The sST2 levels was slightly increased in HTLV-1 patients, though such increase was not statistically significant (P = 0.91), in addition sST2 level did not correlate significantly to the disease duration (P = 0.78). Despite some other chronic viral infection, HTLV-1 seems not to induce high serum sST2. However owing to relatively high normal variation of sST2 levels and rather small sample size, we stongly recommend further reseach with preferably larger sample size to evalute sST2 in HTLV-1 infected patients.

Mechanism of inhibition of ribonucleotide reductase with motexafin gadolinium (MGd).

Motexafin gadolinium (MGd) is an expanded porphyrin anticancer agent which selectively targets tumor cells and works as a radiation enhancer, with promising results in clinical trials. Its mechanism of action is oxidation of intracellular reducing molecules and acting as a direct inhibitor of mammalian ribonucleotide reductase (RNR). This paper focuses on the mechanism of inhibition of RNR by MGd. Our experimental data present at least two pathways for inhibition of RNR; one precluding subunits oligomerization and the other direct inhibition of the large catalytic subunit of the enzyme. Co-localization of MGd and RNR in the cytoplasm particularly in the S-phase may account for its inhibitory properties. These data can elucidate an important effect of MGd on the cancer cells with overproduction of RNR and its efficacy as an anticancer agent and not only as a general radiosensitizer.

Molecular mechanisms of thioredoxin and glutaredoxin as hydrogen donors for Mammalian s phase ribonucleotide reductase.

Ribonucleotide reductase (RNR) catalyzes the rate-limiting step in deoxyribonucleotide synthesis essential for DNA replication and repair. RNR in S phase mammalian cells comprises a weak cytosolic complex of the catalytic R1 protein containing redox active cysteine residues and the R2 protein harboring the tyrosine free radical. Each enzyme turnover generates a disulfide in the active site of R1, which is reduced by C-terminally located shuttle dithiols leaving a disulfide to be reduced. Electrons for reduction come ultimately from NADPH via thioredoxin reductase and thioredoxin (Trx) or glutathione reductase, glutathione, and glutaredoxin (Grx), but the mechanism has not been clarified for mammalian RNR. Using recombinant mouse RNR, we found that Trx1 and Grx1 had similar catalytic efficiency (k(cat)/K(m)). With 4 mm GSH, Grx1 showed a higher affinity (apparent K(m) value, 0.18 microm) compared with Trx1 which displayed a higher apparent k(cat), suggesting its major role in S phase DNA replication. Surprisingly, Grx activity was strongly dependent on GSH concentrations (apparent K(m) value, 3 mm) and a Grx2 C40S mutant was active despite only one cysteine residue in the active site. This demonstrates a GSH-mixed disulfide mechanism for glutaredoxin catalysis in contrast to the dithiol mechanism for thioredoxin. This may be an advantage with the low levels of RNR for DNA repair or in tumor cells with high RNR and no or low Trx expression. Our results demonstrate mechanistic differences between the mammalian and canonical Escherichia coli RNR enzymes, which may offer an explanation for the nonconserved shuttle dithiol sequences in the C terminus of the R1.

Motexafin gadolinium, a tumor-selective drug targeting thioredoxin reductase and ribonucleotide reductase.

Motexafin gadolinium (MGd) is a chemotherapeutic drug that selectively targets tumor cells and mediates redox reactions generating reactive oxygen species. Thioredoxin (Trx), NADPH, and thioredoxin reductase (TrxR) of the cytosol/nucleus or mitochondria are major thiol-dependent reductases with many functions in cell growth, defense against oxidative stress, and apoptosis. Mammalian TrxRs are selenocysteine-containing flavoenzymes; MGd was an NADPH-oxidizing substrate for human or rat TrxR1 with a Km value of 8.65 microM (kcat/Km of 4.86 x 10(4) M(-1) s(-1)). The reaction involved redox cycling of MGd by oxygen producing superoxide and hydrogen peroxide. MGd acted as a non-competitive inhibitor (IC50 of 6 microM) for rat TrxR. In contrast, direct reaction between MGd and reduced human Trx was negligible. The corresponding reaction with reduced Escherichia coli Trx was also negligible, but MGd was a better substrate (kcat/Km of 2.23 x 10(5) M(-1) s(-1)) for TrxR from E. coli and a strong inhibitor of Trx-dependent protein disulfide reduction. Ribonucleotide reductase (RNR), a 1:1 complex of the non-identical R1- and R2-subunits, catalyzes the essential de novo synthesis of deoxyribonucleotides for DNA synthesis using electrons from Trx and TrxR. MGd inhibited recombinant mouse RNR activity with either 3 microM reduced human Trx (IC50 2 microM) or 4 mM dithiothreitol (IC50 6 microM) as electron donors. Our results demonstrate MGd-induced enzymatic generation of reactive oxygen species by TrxR plus a powerful inhibition of RNR. This may explain the effects of the drug on cancer cells, which often overproduce TrxR and have induced RNR for replication and repair.