DDX43 mRNA expression and protein levels in relation to clinicopathological profile of breast cancer.

BACKGROUND: Breast cancer (BC) is the most often diagnosed cancer in women globally. Cancer cells appear to rely heavily on RNA helicases. DDX43 is one of DEAD- box RNA helicase family members. But, the relationship between clinicopathological, prognostic significance in different BC subtypes and DDX43 expression remains unclear. Therefore, the purpose of this study was to assess the clinicopathological significance of DDX43 protein and mRNA expression in different BC subtypes. MATERIALS AND METHODS: A total of 80 females newly diagnosed with BC and 20 control females that were age-matched were recruited for this study. DDX43 protein levels were measured by ELISA technique. We used a real-time polymerase chain reaction quantification (real-time PCR) to measure the levels of DDX43 mRNA expression. Levels of DDX43 protein and mRNA expression within BC patients had been compared to those of control subjects and correlated with clinicopathological data. RESULTS: The mean normalized serum levels of DDX43 protein were slightly higher in control than in both benign and malignant groups, but this result was non-significant. The mean normalized level of DDX43 mRNA expression was higher in the control than in both benign and malignant cases, although the results were not statistically significant and marginally significant, respectively. Moreover, the mean normalized level of DDX43 mRNA expression was significantly higher in benign than in malignant cases. In malignant cases, low DDX43 protein expression was linked to higher nuclear grade and invasive duct carcinoma (IDC), whereas high mRNA expression was linked to the aggressive types of breast cancer such as TNBC, higher tumor and nuclear grades. CONCLUSION: This study explored the potential of using blood DDX43 mRNA expression or protein levels, or both in clinical settings as a marker of disease progression in human breast cancer. DDX43 mRNA expression proposes a less invasive method for discriminating benign from malignant BC.

Surfactant nanovesicles for augmented antibacterial activity against carbapenemase resistant enterobacteriaceae and extended spectrum beta-lactamases producing bacteria: in vitro and in vivo evaluation.

The ubiquitous emergence of bacterial resistance is a challenging problem in infectious diseases treatment. Recently, new research lines employed nano-drug delivery systems to enhance antibacterial activity of the existing antibiotics. Accordingly, the objective of this study is to optimize surfactant nanovesicles to improve the antimicrobial effect of meropenem, ertapenem and tigecycline against Carbapenemase Resistant Enterobacteriaceae (CRE) and extended spectrum beta-lactamases producing bacteria (ESBL). Klebsiella pneumoniae and Escherichia coli were used as the test organisms. In vivo and in vitro evaluations were conducted to prove the efficacy of niosome-encapsulated drugs formulations. The results revealed that surfactant vesicles were able to reduce the MIC values of the tested drugs by nine-fold change compared to their free forms. Scanning Electron Microscope (SEM) showed possible adhesion/fusion of the vesicles encapsulated drugs on the bacterial cells compared to its solution. In vivo investigations using animal skin model confirmed the superiority of nanovesicles drug encapsulation regarding both wound size and histopathological examination. Wound surface area was reduced from 24.6mm2 in absence of drug to reach 13.9, and 6.2mm2 in presence of ertapenem solution or niosomes, respectively. Nanovesicular formulations can be considered as effective drug delivery systems that can diminish bacterial resistance against ß-lactams antibiotics.

Mutational pattern of PIK3CA exon 20 in circulating DNA in breast cancer.

Breast cancer (BC) is one of the most common cancers with diverse mutations, etiology and causes. Mutational signature of the driver genes could allow for better understanding disease etiology and progression. This study aims to assess PIK3CA Exon 20 somatic mutational signature in relation to potential underlying etiology. Circulating DNA of 71 Egyptian BC patients was isolated, amplified for PIK3CA Exon 20, and sequenced. Mutational signature was determined according to COSMIC v2 signature. Public BC dataset was analysed to assess PIK3CA mutations effect on the transcriptomic profile. Somatic mutations of PIK3CA exon 20 were found in 66.2% of the study cohort. Nucleotide substitution patterns were similar to general nucleotide substitution patterns in BC. Signature 3 and 9 were the most common signatures in the studied BC patients. Signature of Aristolochic acid exposure was found in some cases. The most common nucleotide substitution was T > A transversion, but substitutions T > G and T > C were correlated to each other and to the total mutation number. PIK3CA mutations were found to disrupt several pathways including RAC1, PDGF, Wnt, and integrin signalling. PIK3CA exon 20 mutational signatures in Egyptian BC patients could suggest a disease etiology involving homologous recombination deficiency (HRD) and polymerase eta (Pol η). Nucleotide substitution patterns could indicate the role of exposure to oxidative stress and some carcinogens such as 4-aminobiphenyl and Aristolochic acid.

Changes in the R-region interactions depend on phosphorylation and contribute to PKA and PKC regulation of the cystic fibrosis transmembrane conductance regulator chloride channel.

The CFTR chloride channel is regulated by phosphorylation at PKA and PKC consensus sites within its regulatory region (R-region) through a mechanism, which is still not completely understood. We used a split-CFTR construct expressing the N-term-TMD1-NBD1 (Front Half; FH), TMD2-NBD2-C-Term (Back Half; BH), and the R-region as separate polypeptides (Split-R) in BHK cells, to investigate in situ how different phosphorylation conditions affect the R-region interactions with other parts of the protein. In proximity ligation assays, we studied the formation of complexes between the R-region and each half of the Split-CFTR. We found that at basal conditions, the density of complexes formed between the R-region and both halves of the split channel were equal. PKC stimulation alone had no effect, whereas PKA stimulation induced the formation of more complexes between the R-region and both halves compared to basal conditions. Moreover, PKC + PKA stimulation further enhanced the formation of FH-R complexes by 40% from PKA level. In cells expressing the Split-R with the two inhibitory PKC sites on the R-region inactivated (SR-S641A/T682A), density of FH-R complexes was much higher than in Split-R WT expressing cells after PKC or PKC + PKA stimulation. No differences were observed for BH-R complexes measured at all phosphorylation conditions. Since full-length CFTR channels display large functional responses to PKC + PKA in WT and S641A/T682A mutant, we conclude that FH-R interactions are important for CFTR function. Inactivation of consensus PKC site serine 686 (S686A) significantly reduced the basal BH-R interaction and prevented the PKC enhancing effect on CFTR function and FH-R interaction. The phospho-mimetic mutation (S686D) restored basal BH-R interaction and the PKC enhancing effect on CFTR function with enhanced FH-R interaction. As the channel function is mainly stimulated by PKA phosphorylation of the R-region, and this response is known to be enhanced by PKC phosphorylation, our data support a model in which the regulation of CFTR activation results from increased interactions of the R-region with the N-term-TMD1-NBD1. Also, serine S686 was found to be critical for the PKC enhancing effect which requires a permissive BH-R interaction at basal level and increased FH-R interaction after PKC + PKA phosphorylation.

Association of Serum Cholesterol Ester Transfer Protein Levels with Taq IB Polymorphism in Acute Coronary Syndrome.

Information on the relationship between circulating cholesteryl ester transfer protein (CETP) levels and coronary heart disease (CHD) incidence (and also, therefore, acute coronary syndrome [ACS]) is conflicting. Many studies have been published concerning this relationship, most of which have incompatible results. In our study, we aimed to determine serum CETP levels in subject individuals with ACS and healthy control individuals, and the association of those levels with Taq IB polymorphism. The current study was conducted with 62 hospitalized patients who had been diagnosed with ACS and 26 controls. All subjects were selected from a previous study of which we are among the coauthors. Serum CETP levels were determined by quantitative enzyme-linked immunosorbent assay (ELISA). The mean serum CETP levels in all patients were significantly higher than those in controls. CETP TaqIB polymorphism affected serum CETP levels, with higher serum CETP for the GA genotype in both groups than in other genotypes. Although the AA genotype showed higher CETP levels than the GG genotype in patients with ACS, the GG showed higher CETP than the AA in healthy controls. Our results support an association between high serum CETP and ACS incidence. Our study helped address some of the controversies regarding the relationship of serum CETP mass to atherosclerosis, in addition to the association of ACS occurrence with circulating CETP levels.

Association of Cholesterol Ester Transfer Protein Taq IB Polymorphism With Acute Coronary Syndrome in Egyptian National Patients.

BACKGROUND: The association between cholesterol ester transfer protein (CETP) Taq IB polymorphism and coronary artery disease (CAD) has been studied in different populations. Acute coronary syndrome (ACS) is a group of clinical symptoms within acute myocardial ischemia, including unstable angina (UA) and myocardial infarction (MI). Because there are no data reported in the literature concerning the cholesteryl ester transfer protein (CETP) Taq IB polymorphism in Egyptians, our study aimed to investigate the frequency of different CETP Taq IB genotypes in Egyptian patients with ACS and in healthy control individuals. METHODS: The current study was conducted with 70 hospitalized patients who had been diagnosed with ACS and 30 controls. We used real-time polymerase chain reaction (RT-PCR) to determine CETP Taq IB in individuals with different genotypes. RESULTS: The frequency of the GA genotype was significantly lower in UA patients, compared with the control group ( P <.05). CONCLUSIONS: The frequency of the CETP Taq IB genotypes and alleles in all groups was similar to that in other ethnic groups. Individuals with the Taq IB GA genotype may have a lower risk of UA.

Factor structure and reliability of the Arabic adaptation of the Hypomania Check List-32, second revision (HCL-32-R2).

OBJECTIVE: To assess the psychometric properties of the Arabic adaptation of the Hypomania-Check-List 32-item, second revision (HCL-32-R2) for the detection of bipolarity in major depressive disorder (MDD) inpatients suffering a current major depressive episode (MDE). METHOD: The "Bipolar Disorders: Improving Diagnosis, Guidance, and Education" Arabic module of the HCL-32-R2 was administered to mother-tongue Arabic MDE inpatients between March 2013 and October 2014. Diagnostic and Statistical Manual Fourth edition (DSM-IV) diagnoses were made adopting the mini-international neuropsychiatric interview, using bipolar disorder (BD) patients as controls. RESULTS: In our sample (n=500, of whom, BD-I=329; BD-II=70; MDD=101), using a cut-off of 17 allowed the HCL-32-R2 to discriminate DSM-IV-defined MDD patients between "true unipolar" (HCL-32-R2(-)) and "sub-threshold bipolar depression" (HCL-32-R2(+)) with sensitivity=82% and specificity=77%. Area under the curve was .883; positive and negative predictive values were 93.44% and 73.23% respectively. Owing to clinical interpretability considerations and consistency with previous adaptations of the HCL-32, a two-factor solution (F1="hyperactive/elated" vs. F2="irritable/distractible/impulsive") was preferred using exploratory and confirmatory factors analyses. Item n.33 ("I gamble more") and n.34 ("I eat more") introduced in the R2 version of the HCL-32 loaded onto F1, though very slightly. Cronbach's alphas were F1=.86 and F2=.60. LIMITATIONS: No cross-validation with any additional validated screening tool. Inpatients only sample; recall bias; no systematic evaluation of eventual medical/psychiatric comorbidities, current/lifetime pharmacological history, or record of severity of current MDE. CONCLUSIONS: In our sample, the HCL-32 fairly discriminated between MDD and BD-I but not BD-II, therefore soliciting for replication studies for use in Arabic-speaking depressed inpatients.

PKC phosphorylation modulates PKA-dependent binding of the R domain to other domains of CFTR.

Activity of the CFTR channel is regulated by phosphorylation of its regulatory domain (RD). In a previous study, we developed a bicistronic construct called DeltaR-Split CFTR, which encodes the front and back halves of CFTR as separate polypeptides without the RD. These fragments assemble to form a constitutively active CFTR channel. Coexpression of the third fragment corresponding to the missing RD restores regulation by PKA, and this is associated with dramatically enhanced binding of the phosphorylated RD. In the present study, we examined the effect of PKC phosphorylation on this PKA-induced interaction. We report here that PKC alone enhanced association of the RD with DeltaR-Split CFTR and that binding was further enhanced when the RD was phosphorylated by both kinases. Mutation of all seven PKC consensus sequences on the RD (7CA-RD) did not affect its association under basal (unphosphorylated) conditions but abolished phosphorylation-induced binding by both kinases. Iodide efflux responses provided further support for the essential role of RD binding in channel regulation. The basal activity of DeltaR-Split/7CA-RD channels was similar to that of DeltaR-Split/wild type (WT)-RD channels, whereas cAMP-stimulated iodide efflux was greatly diminished by removal of the PKC sites, indicating that 7CA-RD binding maintains channels in an inactive state that is unresponsive to PKA. These results suggest a novel mechanism for CFTR regulation in which PKC modulates PKA-induced domain-domain interactions.