Elrasheid A. H. Kheirelseid

MicroRNA signature analysis in colorectal cancer: identification of expression profiles in stage II tumors associated with aggressive disease

PurposeColorectal cancer (CRC) is a clinically diverse disease whose molecular etiology remains poorly understood. The purpose of this study was to identify miRNA expression patterns predictive of CRC tumor status and to investigate associations between microRNA (miRNA) expression and clinicopathological parameters.MethodsExpression profiling of 380 miRNAs was performed on 20 paired stage II tumor and normal tissues. Artificial neural network (ANN) analysis was applied to identify miRNAs predictive of tumor status. The validation of specific miRNAs was performed on 102 tissue specimens of varying stages.ResultsThirty-three miRNAs were identified as differentially expressed in tumor versus normal tissues. ANN analysis identified three miRNAs (miR-139-5p, miR-31, and miR-17-92 cluster) predictive of tumor status in stage II disease. Elevated expression of miR-31 (p = 0.004) and miR-139-5p (p < 0.001) and reduced expression of miR-143 (p = 0.016) were associated with aggressive mucinous phenotype. Increased expression of miR-10b was also associated with mucinous tumors (p = 0.004). Furthermore, progressively increasing levels of miR-10b expression were observed from T1 to T4 lesions and from stage I to IV disease.ConclusionAssociation of specific miRNAs with clinicopathological features indicates their biological relevance and highlights the power of ANN to reliably predict clinically relevant miRNA biomarkers, which it is hoped will better stratify patients to guide adjuvant therapy.

Identification of endogenous control genes for normalisation of real-time quantitative PCR data in colorectal cancer

BackgroundGene expression analysis has many applications in cancer diagnosis, prognosis and therapeutic care. Relative quantification is the most widely adopted approach whereby quantification of gene expression is normalised relative to an endogenously expressed control (EC) gene. Central to the reliable determination of gene expression is the choice of control gene. The purpose of this study was to evaluate a panel of candidate EC genes from which to identify the most stably expressed gene(s) to normalise RQ-PCR data derived from primary colorectal cancer tissue.ResultsThe expression of thirteen candidate EC genes: B2M, HPRT, GAPDH, ACTB, PPIA, HCRT, SLC25A23, DTX3, APOC4, RTDR1, KRTAP12-3, CHRNB4 and MRPL19 were analysed in a cohort of 64 colorectal tumours and tumour associated normal specimens. CXCL12, FABP1, MUC2 and PDCD4 genes were chosen as target genes against which a comparison of the effect of each EC gene on gene expression could be determined. Data analysis using descriptive statistics, geNorm, NormFinder and qBasePlus indicated significant difference in variances between candidate EC genes. We determined that two genes were required for optimal normalisation and identified B2M and PPIA as the most stably expressed and reliable EC genes.ConclusionThis study identified that the combination of two EC genes (B2M and PPIA) more accurately normalised RQ-PCR data in colorectal tissue. Although these control genes might not be optimal for use in other cancer studies, the approach described herein could serve as a template for the identification of valid ECs in other cancer types.

Younger age as a prognostic indicator in breast cancer: A cohort study

BackgroundThe debate continues as to whether younger women who present with breast cancer have a more aggressive form of disease and a worse prognosis. The objectives of this study were to determine the incidence of breast cancer in women under 40 years old and to analyse the clinicopathological characteristics and outcome compared to an older patient cohort.MethodsData was acquired from a review of charts and the prospectively reviewed GUH Department of Surgery database. Included in the study were 276 women diagnosed with breast cancer under the age of forty and 2869 women over forty. For survival analysis each women less than 40 was matched with two women over forty for both disease stage and grade.ResultsThe proportion of women diagnosed with breast cancer under the age of forty in our cohort was 8.8%. In comparison to their older counterparts, those under forty had a higher tumour grade (p = 0.044) and stage (p = 0.046), a lower incidence of lobular tumours (p < 0.001), higher estrogen receptor negativity (p < 0.001) and higher HER2 over-expression (p = 0.002); there was no statistical difference as regards tumour size (p = 0.477). There was no significant difference in overall survival (OS) for both groups; and factors like tumour size (p = 0.026), invasion (p = 0.026) and histological type (p = 0.027), PR (p = 0.031) and HER2 (p = 0.002) status and treatment received were independent predictors of OSConclusionBreast cancer in younger women has distinct histopathological characteristics; however, this does not result in a reduced survival in this population.

A comparative study of the bell-bottom technique vs hypogastric exclusion for the treatment of aneurysmal extension to the iliac bifurcation

INTRODUCTION A significant proportion of patients undergoing endovascular aneurysm repair (EVAR) have common iliac artery aneurysms (CIAA). Aneurysmal involvement at the iliac bifurcation potentially undermines long-term durability. METHODS Patients with CIAA who underwent EVAR were identified in two teaching hospitals. Bell-bottom technique (BBT; iliac limb ≥20 mm) or internal iliac artery embolization and limb extension to the external iliac artery (IIE + EE) were used. Outcome between these two approaches was compared. RESULTS We identified 185 patients. Indication for EVAR included asymptomatic abdominal aortic aneurysm (AAA) in 157, symptomatic or ruptured aneurysm in 19, and CIAA in nine. Mean AAA diameter was 59 mm. Among 260 large CIAAs that were treated, BBT was used to treat 166 CIAA limbs, and 94 limbs underwent IIE + EE. Total reintervention rates were 11% for BBT (n = 19) and 19.1% for IIE + EE (n = 18; P = .149). Rates of reintervention for type Ib or III endoleak were 4% for BBT (n = 7) and 4% for IIE + EE (n = 4; P > .99). The difference in limb patency rates was not significant. The 30-day mortality rate was 1%. Median follow-up was 22 months. Complications did not differ significantly between the two groups; however, the combined incidence of perioperative complications and reinterventions was higher in the IIE + EE group (49% vs 22%; P = .002). CONCLUSIONS The combined incidence of perioperative complications and reinterventions is significantly higher with IIE + EE than with BBT; therefore, when feasible, BBT is desirable.

Clinical applications of gene expression in colorectal cancer

BACKGROUND Despite developments in diagnosis and treatment, 20% of colorectal cancer (CRC) patients present with metastatic disease and 30% of cases recur after curative surgery. Furthermore, the molecular factors involved in prognosis and response to therapy in CRC is poorly understood. The aims of this study were to quantitatively examine the expression of target genes in colorectal cancer and to correlate their expression levels with clinico-pathological variables. METHODS A detailed analysis of published CRC microarray data was performed to identify the most prominent genes. The selected genes were validated in fifty-two pairs of fresh colorectal tumour and associated normal tissue specimens by RQ-PCR using TaqMan(®) assays. Statistical analysis and correlation with clinicopathological data was performed using SPSS software. RESULTS Expression levels of CXCL12 (P=0.000), CDH17 (P=0.026), MUC2 (P=0.000), L-FABP (P=0.000) and PDCD4 (P=0.000) were down regulated and IL8 (P=0.000) was upregulated in tumours compared to normal colorectal tissues. No significant differences were noted in expression of CEACAM5, CXCR4, CXCR7, TGFB1, TGFBR1 and TGFBR2. Furthermore, we found significant associations of gene expression levels and clinicopathological variables such as tumour size, grade, invasion and lymph node status. CONCLUSIONS We identified a comprehensive list of genes with highly differential expression patterns in colorectal cancer that could serve as molecular markers to complement existing histopathological factors in diagnosis, follow up and therapeutic strategies for individualised care of patients.

Role of Far Infra-Red Therapy in Dialysis Arterio-Venous Fistula Maturation and Survival: Systematic Review and Meta-Analysis

Introduction A well-functioning arteriovenous fistula (AVF) is the best modality for vascular access in patients with end-stage renal disease (ESRD) requiring haemodialysis (HD). However, AVFs’ main disadvantage is the high rate of maturation failure, with approximately one third (20%–50%) not maturing into useful access. This review examine the use of Far-Infra Red therapy in an attempt to enhance both primary (unassisted) and secondary (assisted) patency rates for AVF in dialysis and pre-dialysis patients. Method We performed an online search for observational studies and randomised controlled trials (RCTs) that evaluated FIR in patients with AVF. Eligible studies compared FIR with control treatment and reported at least one outcome measure relating to access survival. Primary patency and secondary patency rates were the main outcomes of interest. Results Four RCTs (666 patients) were included. Unassisted patency assessed in 610 patients, and was significantly better among those who received FIR (228/311) compared to (185/299) controls (pooled risk ratio of 1.23 [1.12–1.35], p = 0.00001). In addition, the two studies which reported secondary patency rates showed significant difference in favour of FIR therapy- 160/168 patients - compared to 140/163 controls (pooled risk ratio of 1.11 [1.04–1.19], p = 0.003). Conclusion FIR therapy may positively influence the complex process of AVF maturation improving both primary and secondary patency rates. However blinded RCTs performed by investigators with no commercial ties to FIR therapy technologies are needed.

Mismatch repair protein expression in colorectal cancer

INTRODUCTION Alterations in at least six of the genes that encode proteins involved in the mismatch repair (MMR) system have been identified in either HNPCC or sporadic colon cancer. We aimed to analyse the proportion of patients with colorectal cancer with loss of immunostaining for MMR proteins in order to determine the feasibility of molecular screening for the loss of MMR proteins through the study of unselected patients with colorectal cancer. METHODS A group of 33 patients with colorectal cancer was randomly selected from the department of surgery bio-bank to determine the expression of MMR proteins in their FFPE tumour tissues using immunohistochemistry techniques. Changes in protein expression following transfection of colorectal tissues were observed in stained cells using Olympus BX60 microscope and image analySIS software. RESULTS Of the tissue specimens in which acceptable immunostaining was achieved, three samples showed loss of one or more of the MMR proteins. Both hMLH1 and hPMS2 proteins were not expressed in a 36 years old woman with cancer of the caecum. The expression of hMSH6 protein was undetermined in tumour tissues retrieved from a 61 years old man with cancer of the proximal colon. The third case was a 77 years old man with no documented family history of cancer, who had carcinoma of the rectum. He showed loss of hMLH1 expression in the tumour tissues. CONCLUSIONS Our findings and the previous reports pointed out the importance of molecular screening of patients with colorectal cancer for MSI using immunohistochemistry. This strategy managed to identify mutations in patients otherwise would not have been detected.

Comparing Supervised Exercise Therapy to Invasive Measures in the Management of Symptomatic Peripheral Arterial Disease

Peripheral arterial disease (PAD) is associated with considerable morbidity and mortality. Consensus rightly demands the incorporation of supervised exercise training (SET) into PAD treatment protocols. However, the exact role of SET particularly its relationship with intervention requires further clarification. While supervised exercise is undoubtedly an excellent tool in the conservative management of mild PAD its use in more advanced disease as an adjunct to open or endovascular intervention is not clearly defined. Indeed its use in isolation in this cohort is incompletely reported. The aim of this review is to clarify the exact role of SET in the management of symptomatic PAD and in particular to assess its role in comparison with or as an adjunct to invasive intervention. A systematic literature search revealed a total 11 randomised studies inclusive of 969 patients. All studies compared SET and intervention with monotherapy. Study results suggest that exercise is a complication-free treatment. Furthermore, it appears to offer significant improvements in patients walk distances with a combination of both SET and intervention offering a superior walking outcome to monotherapy in those requiring invasive measures.

mRNA/miRNA correlations in colorectal cancer: novel mechanisms in cancer initiation and progression

miRNAs are small noncoding RNAwhich are cleaved from 70 to 100 nt endogenous double-stranded precursors. Although their biological role is not fully understood, miRNAs are found in diverse organisms and epigenetically function as negative regulators of gene expression. They are complementary to genomic regions, and one of their modes of action is to bind three prime untranslated region (3′-UTR) and inhibit gene translation. Moreover, they can also function by cleaving and degrading a target mRNA, in which case the miRNA may target sequences outside the 3′-UTR. Functional characterisation of miRNAs will depend heavily on the identification of their specific gene targets. In addition, a number of studies have shown that more than one miRNA can potentially bind to a single targeted gene; hence, multiple miRNAs may cooperatively control the expression of target genes. Numerous bioinformatic methods were developed to high-throughput prediction of miRNA target genes, although it is understood that the presumed targets have to be validated experimentally. Computational approaches have been developed based on an understanding of the relationship between the miRNA seed region and the 3′-UTR of the target gene. To develop computational algorithms, empirical evidence is examined carefully and principles of miRNA target recognition are extracted. After preparation of the data set, miRNA binding sites are identified by determining the base pairing pattern of miRNAs and mRNAs according to the complementarity within specific region, thermodynamic analysis of the miRNA/mRNA duplexes via calculation of the free energy and comparative sequence analysis. Then, the number of target sites of miRNA is counted. However, most of the available computational prediction algorithms have a group of drawbacks which can result in the prediction of false positives. Moreover, some targets may pass undetected. The false positive rates were estimated at 22, 24 and 30 % for TargetScan, miRanda and PicTar, respectively [1]. The PicTar and EMBL algorithms have a reported sensitivity of 70–80 % [2] indicating that 20–30 % of targets may go undetected. Therefore, it is necessary to confirm the bioinformatically presumed miRNA target experimentally. Verification of the biological function or target gene of a miRNA can be performed using gain-of-function and loss-offunction approaches in vitro and in vivo. As miRNAs modulate gene expression by both mRNA degradation and Electronic supplementary material The online version of this article (doi:10.1007/s00384-012-1574-8) contains supplementary material, which is available to authorized users. E. A. H. Kheirelseid (*) :N. Miller :K. H. Chang :M. J. Kerin Department of Surgery, National University of Ireland Galway, Clinical Science Institute, Costello Road, Galway, Ireland e-mail: rashmed1111@gmail.com

Supervised exercise therapy in the management of peripheral arterial disease – an assessment of compliance

BACKGROUND Supervised exercise therapy (SET) is an effective option in the management of peripheral arterial disease (PAD). Unfortunately, poor compliance remains prevalent. This study aimed to assess patient exercise compliance and to identify factors influencing symptomatic improvement and SET participation. PATIENTS AND METHODS Data regarding attendance at SET for this cohort study were extracted from a prospectively maintained database of patients with claudication attending SET at Dublin City University. All patients had ankle brachial index confirmed PAD with associated intermittent claudication. Exercise performance and symptomatic data were gathered retrospectively using patient charts and interviews. RESULTS Ninety-eight patients were referred for SET during the study period. The mean age was 69.2 (± 10.1) with 18 % being female. Median follow-up was 25.1 months (IQ range 17.0-31.6). Overall, the mean number of sessions attended per patient was 19.5. Exercise compliance was associated with a significant improvement in symptoms (p = 0.001). Other factors including anatomical level of claudication (P = 0.042) and educational level (p = 0.007) were found to affect the outcome of SET. Multivariate analysis revealed hypertension as a predictor of symptomatic outcome after SET (p = 0.045). Furthermore, ex-smokers (p = 0.021) and those previously diagnosed with hypercholesterolaemia (p = 0.020) or ischaemic heart disease (p = 0.029) had superior exercise compliance. Using linear regression, smoking history (p = 0.024) was identified as a predictor of compliance to SET. CONCLUSIONS Establishing exercise compliance remains challenging in the PAD cohort. Pre-exercise patient education and personalised exercise prescriptions may result in improvements in function and compliance.