MicroRNAs in cancer drug resistance: Basic evidence and clinical applications

Development of drug resistance has considerably limited the efficacy of cancer treatments, including chemotherapy and targeted therapies. Hence, understanding the molecular mechanisms underpinning the innate or the acquired resistance to these therapies is critical to improve drug efficiency and clinical outcomes. Several studies have implicated microRNAs (miRNA) in this process. MiRNAs repress gene expression by specific binding to complementary sequences in the 3' region of target messenger RNAs (mRNAs), followed by target mRNA degradation or blocked translation. By targeting molecules specific to a particular pathway within tumor cells, the new generation of cancer treatment strategies has shown significant advantages over conventional chemotherapy. However, the long-term efficacy of targeted therapies often remains poor, because tumor cells develop resistance to such therapeutics. Targeted therapies often involve monoclonal antibodies (mAbs), such as those blocking the ErB/HER tyrosine kinases, epidermal growth factor receptor (cetuximab) and HER2 (trastuzumab), and those inhibiting vascular endothelial growth factor receptor signaling (e.g., bevacizumab). Even though these are among the most used agents in tumor medicine, clinical response to these drugs is reduced due to the emergence of drug resistance as a result of toxic effects in the tumor microenvironment. Research on different types of human cancers has revealed that aberrant expression of miRNAs promotes resistance to the aforementioned drugs. In this study, we review the mechanisms of tumor cell resistance to mAb therapies and the role of miRNAs therein. Emerging treatment strategies combine therapies using innovative miRNA mimics or antagonizers with conventional approaches to maximize outcomes of patients with cancer.     

High-density lipoprotein functionality and breast cancer: A potential therapeutic target

Breast cancer is a major cause of death globally, and particularly in developed countries. Breast cancer is influenced by cholesterol membrane content, by affecting the signaling pathways modulating cell growth, adherence, and migration. Furthermore, steroid hormones are derived from cholesterol and these play a key role in the pathogenesis of breast cancer. Although most findings have reported an inverse association between serum high-density lipoprotein (HDL)-cholesterol level and the risk of breast cancer, there have been some reports of the opposite, and the association therefore remains unclear. HDL is principally known for participating in reverse cholesterol transport and has an inverse relationship with the cardiovascular risk. HDL is heterogeneous, with particles varying in composition, size, and structure, which can be altered under different circumstances, such as inflammation, aging, and certain diseases. It has also been proposed that HDL functionality might have a bearing on the breast cancer. Owing to the potential role of cholesterol in cancer, its reduction using statins, and particularly as an adjuvant during chemotherapy may be useful in the anticancer treatment, and may also be related to the decline in cancer mortality. Reconstituted HDLs have the ability to release chemotherapeutic drugs inside the cell. As a consequence, this may be a novel way to improve therapeutic targeting for the breast cancer on the basis of detrimental impacts of oxidized HDL on cancer development.     

Comparison of five molecular subtyping methods for differentiation of Salmonella Kentucky isolates in Tunisia

Salmonellosis is one of the most common causes of food-borne infection worldwide. In the last decade, Salmonella enterica serovar Kentucky has shown an increase in different parts of the world with the concurrent emergence of multidrug-resistant isolates. These drug-resistant types spread from Africa and the Middle East to Europe and Asia. Although S. Kentucky serovar is of potential human relevance, there is currently no standardized fingerprinting method for it, in Tunisia. In the present study, a collection of 57 Salmonella Kentucky isolates were analyzed using plasmid profiling, pulsed-field gel electrophoresis (PFGE), ribotyping, enterobacterial repetitive intergenic consensus (ERIC) fingerprinting, and Random Amplification of Polymorphic DNA. Plasmid profiling showed a discriminatory index (D) of 0.290, and only 9 out of 57 (16 %) isolates carried plasmids, which represents a limitation of this technique. Fingerprinting of genomic DNA by PFGE and ribotyping produced 4 and 5 patterns, respectively. Distinct PFGE patterns (SX1, SX2, SX3, and SX4) were generated for only 28 strains out of 57 (49.1 %) with a D value of 0.647. RAPD fingerprinting with primers RAPD1 and RAPD2 produced 4 and 20 patterns, respectively. ERIC fingerprinting revealed 14 different patterns with a high discriminatory index (D) of 0.903. When the methods were combined, the best combination of two methods was ERIC-2 with RAPD2. These results indicates that a single method cannot be relied upon for discriminating between S. Kentucky strains, and a combination of typing methods such ERIC2 and RAPD2 allows further discrimination.     

Renin-angiotensin system polymorphisms in relation to hypertension status and obesity in a Tunisian population

Essential hypertension (HTA) is the clinical expression of a disordered interaction between the genetic, physiological, and biochemical systems that under usual conditions maintain cardiovascular homeostasis. We studied the effects of the angiotensinogen M235T, angiotensin converting enzyme insertion/deletion (ACE I/D), and angiotensin II receptor 1 (AT1R) A1166C gene polymorphisms on the risk of HTA and to evaluate the relationship between these polymorphisms and obesity. We performed AGT, ACE and AGTR genotyping in 142 hypertensive patients and 191 control subjects using PCR-RFLP methods and PCR, respectively. The three polymorphisms were significantly associated with HTA. Individuals carrying the mutated TT of AGT, DD of ACE and CC of AT1R genotypes had an 1.67 (P = 0.032), 3.09 (P < 0.001) and 3.45 (P < 0.001)-fold increased risk of HTA. After adjustment for sex, smoking, diabetes, dyslipidemia, BMI, triglycerides and DD, TT and CC genotypes, BMI was independent risk factor of HTA (OR = 3.14; P < 0.001). An association of BMI with ACE gene polymorphism (P = 0.035), whereas no association with AGT and AT1R gene polymorphisms was obtained. The proportion of hypertensives is as high as 21.8 and 13.4% in the overweight and the obese DD group. The present study implies that the genotyping for the variants of RAS gene could in the future become an important part of the clinical process of risk identification for HTA.     

The beneficial effect of repaglinide on in vitro maturation and development ability of immature mouse oocytes

Repaglinide is a hypoglycemic drug, causing depolarization of the cell membrane, opening the voltage-gated calcium channels, and then increasing intracellular calcium in the pancreatic B cells by inhibition of the K-ATP-sensitive channels. Oocyte in vitro maturation (IVM) is influenced by different factors such as calcium signaling. In this study, we examined the effects of repaglinide on in vitro maturation and fertilization ability of mouse oocyte. Immature oocytes were isolated from female Naval Medical Research Institute mice which are 6–8 wk old mechanically and then cultured in 30 μl droplets of T6 medium with different concentrations of repaglinide. The control group did not receive repaglinide (R₀). Treatment groups received different concentrations (5, 10, and 100 nM and 1 and 10 μM) of repaglinide (R₁, R₂, R₃, R₄, and R₅, respectively). Oocyte in vitro maturation rate was assessed after 24 h. In vitro fertilization was performed using metaphase II oocytes obtained from R₀ and R₄ treatments. Embryo cleavage rate was calculated at 48 h post-IVF. Chi-square test was used for evaluating difference between control and treatment groups (p < 0.05). Oocyte maturation rate after 24 h in treatment groups R₂, R₃, R₄, and R₅ was significantly higher than that in the control (p < 0.05). Supplementation of medium with 1 μM of repaglinide (R₄) during IVM significantly improved outcome of embryo cleavage rate than control at 48 h post-IVF (p < 0.05). In conclusion, repaglinide can be considered as an effective agent for in vitro oocyte maturation and embryo cleavage.     

The Effects of Synbiotic Supplementation on Pregnancy Outcomes in Gestational Diabetes

Synbiotics are known to exert multiple beneficial effects, including anti-inflammatory and antioxidative actions. This study was designed to evaluate the effects of synbiotic administration on biomarkers of inflammation, oxidative stress, and pregnancy outcomes among gestational diabetic (GDM) women. This randomized, double-blind, placebo-controlled clinical trial was carried out among 60 subjects with GDM who were not on oral hypoglycemic agents. Patients were randomly assigned to consume either one synbiotic capsule containing Lactobacillus acidophilus strain T16 (IBRC-M10785), L. casei strain T2 (IBRC-M10783), and Bifidobacterium bifidum strain T1 (IBRC-M10771) (2 × 10⁹ CFU/g each) plus 800 mg inulin (HPX) (n = 30) or placebo (n = 30) for 6 weeks. Compared with the placebo, synbiotic supplementation significantly decreased serum high-sensitivity C-reactive protein (hs-CRP) (? 1.9 ± 4.2 vs. +1.1 ± 3.5 mg/L, P = 0.004), plasma malondialdehyde (MDA) (? 0.1 ± 0.6 vs. + 0.3 ± 0.7 μmol/L, P = 0.02), and significantly increased total antioxidant capacity (TAC) (+ 70.1 ± 130.9 vs. ? 19.7 ± 124.6 mmol/L, P = 0.009) and total glutathione (GSH) levels (+ 28.7 ± 61.5 vs. ? 14.9 ± 85.3 μmol/L, P = 0.02). Supplementation with synbiotic had a significant decrease in cesarean section rate (16.7 vs. 40.0%, P = 0.04), lower incidence of hyperbilirubinemic newborns (3.3 vs. 30.0%, P = 0.006), and newborns’ hospitalization (3.3 vs. 30.0%, P = 0.006) compared with the placebo. Synbiotic supplementation did not affect plasma nitric oxide (NO) levels and other pregnancy outcomes. Overall, synbiotic supplementation among GDM women for 6 weeks had beneficial effects on serum hs-CRP, plasma TAC, GSH, and MDA; cesarean section; incidence of newborn’s hyperbilirubinemia; and newborns’ hospitalization but did not affect plasma NO levels and other pregnancy outcomes. http://www.irct.ir: www.irct.ir: IRCT201704205623N108     

Proteomics screening of molecular targets of granulocyte colony stimulating factor in the mouse brain and PC12 cell line

Aims

Granulocyte colony stimulating factor (G-CSF), a new neuroprotective agent, binds to its specific receptors in the brain. In this study we hypothesized that at least a part of G-CSF's neuroprotective effect may be mediated through its interaction with other proteins in the brain.

Main methods

Using an immunoprecipitation (IP) kit, at first the antibody of G-CSF was covalently crosslinked to protein A/G agarose. Then the mouse brain or PC12 cell lysate mixed with G-CSF was added to the agarose beads plus antibody. After immunoaffinity isolation of target proteins, gel electrophoresis was performed and protein bands were identified using MALDI-TOF/TOF and MASCOT software.

Key findings

Our data show that G-CSF physically binds to cellular proteins like sodium/potassium-transporting ATPase, beta actin, aldehyde dehydrogenase, regucalcin and glutathione-s-transferase. These proteins are involved in membrane transportation, cell structure, signal transduction, enzymes involve in calcium related cell signaling and redox homeostasis.

Significance

Interaction of G-CSF with these proteins can explain some of its pharmacological effects in the CNS.     

The effects of Nigella sativa (Ns), Anthemis hyalina (Ah) and Citrus sinensis (Cs) extracts on the replication of coronavirus and the expression of TRP genes family

Extracts of Anthemis hyalina (Ah), Nigella sativa (Ns) and peels of Citrus sinensis (Cs) have been used as folk medicine to fight antimicrobial diseases. To evaluate the effect of extracts of Ah, Ns and Cs on the replication of coronavirus (CoV) and on the expression of TRP genes during coronavirus infection, HeLa-CEACAM1a (HeLa-epithelial carcinoembryonic antigen-related cell adhesion molecule 1a) cells were inoculated with MHV-A59 (mouse hepatitis virus–A59) at moi of 30. 1/50 dilution of the extracts was found to be the safe active dose. ELISA kits were used to detect the human IL-8 levels. Total RNA was isolated from the infected cells and cDNA was synthesized. Fluidigm Dynamic Array nanofluidic chip 96.96 was used to analyze the mRNA expression of 21 TRP genes and two control genes. Data was analyzed using the BioMark digital array software. Determinations of relative gene expression values were carried out by using the 2^???Ct method (normalized threshold cycle (Ct) value of sample minus normalized Ct value of control). TCID₅₀/ml (tissue culture infectious dose that will produce cytopathic effect in 50 % of the inoculated tissue culture cells) was found for treatments to determine the viral loads. The inflammatory cytokine IL-8 level was found to increase for both 24 and 48 h time points following Ns extract treatment. TRPA1, TRPC4, TRPM6, TRPM7, TRPM8 and TRPV4 were the genes which expression levels changed significantly after Ah, Ns or Cs extract treatments. The virus load decreased when any of the Ah, Ns or Cs extracts was added to the CoV infected cells with Ah extract treatment leading to undetectable virus load for both 6 and 8hpi. Although all the extract treatments had an effect on IL-8 secretion, TRP gene expression and virus load after CoV infection, it was the Ah extract treatment that showed the biggest difference in virus load. Therefore Ah extract is the best candidate in our hands that contains potential treatment molecule(s).