Stella Marousi

T helper 1 (Th1)/Th2 cytokine expression shift of peripheral blood CD4+ and CD8+ T cells in patients at the post-acute phase of stroke

Local humoral and cellular immune responses modulate the inflammatory processes involved in the development of atherosclerotic lesions, as well as in the evolution of brain infarcts in stroke patients. The role of systemic adaptive immunity on the progression of such disease manifestations is less clear. In the current study, we evaluated the percentages of T helper 1 (Th1) [interleukin (IL)‐2, interferon (IFN)‐γ] and Th2 (IL‐4, IL‐10) cytokine‐producing peripheral blood CD4+ and CD8+ T cells in 23 patients with a history of ischaemic stroke (IS) at the chronic stable phase of the disease (median post‐stroke time 34·5 months). Seven stroke‐free individuals matched for age and vascular risk factors (matched controls, MC) were collected for comparison. To measure cytokine values at baseline and after stimulation, we used a flow cytometry method of intracellular cytokine staining. Intrinsic Th1 and Th2 cytokine production in unstimulated T cells was negligible in all study participants. Following mitogenic stimulation with phorbol 12‐myristate13‐acetate/ionomycin, both the IS and the MC groups exhibited a similarly strong Th1 response; IL‐2 production predominated in the CD4+ T cells and IFN‐γ in the CD8+ T cells. However, when measuring the Th2 cytokine‐production capacity post‐stimulation, a significant increase in the percentage of IL‐4‐producing T cells was observed in the IS groups, compared with the MC group, resulting in a significantly lower ratio of IFN‐γ‐/IL‐4‐producing T cells. No such Th2 enhancement could be confirmed for the case of IL‐10. We propose that in IS patients there is a systemic shift of the immune system towards Th2 responses at the late post‐acute phase of stroke.

The survivin -31 snp in human colorectal cancer correlates with survivin splice variant expression and improved overall survival

Background: Survivin is involved in the regulation of cell division and survival, two key processes in cancer. The majority of studies on survivin in colorectal cancer (CRC) have focused on protein expression and less is known about the expression of survivin splicing variants or survivin gene polymorphisms in CRC. In the present study, the mRNA levels of the five known isoforms of survivin as well as survivin protein were assessed in matched normal and neoplastic colorectal tissue. Moreover, the 9386C/T and −31G/C polymorphisms were investigated. Methods: Quantitative RT-PCR was used to assess mRNA levels in fresh/frozen tissue samples. Protein levels were immunohistochemically evaluated on formalin-fixed paraffin-embedded tissue sections. Individuals were genotyped using real time PCR. Results: Expression of all 5 survivin splice variants as well as survivin protein was elevated in colorectal carcinomas compared to normal tissue. Specific splice variant expression differentially correlated with clinicopathological parameters. Furthermore, both snps correlated with splice variant levels or their ratios in colorectal carcinomas while the −31G/C snp may be related to CRC development and improved overall survival. Conclusion: Our results support a role of survivin in colorectal carcinogenesis while the −31G/C snp may constitute a marker of survival.

Prion protein expression and the M129V polymorphism of the PRNP gene in patients with colorectal cancer.

The prion protein, PrPC, is known mostly for its involvement in neurodegenerative spongiform encephalopathies. However, a role for this molecule in cancer is becoming increasingly recognized partly because it promotes cell proliferation and inhibits apoptosis. Moreover, the codon 129 polymorphism (M129V) of the PRNP gene (the PrPC‐encoding gene) has been associated with neurodegenerative disease development and severity, while no information is available regarding its role in colorectal cancer (CRC) incidence and disease progression. We have previously reported that expression levels of PRNP may have a prognostic value in CRC, suggesting a role for the prion protein in CRC. The aim of this study was to investigate retrospectively the possible role of M129V and PrPC expression in patients with CRC. The M129V single nucleotide polymorphism was genotyped by real time polymerase chain reactions in 110 patients with CRC and 124 healthy donors. Moreover, protein expression was assessed by immunohistochemistry in 68 patients with CRC. Allele frequencies were similar in patients and healthy controls indicating that the M129V polymorphism is not a risk factor for CRC. Furthermore, it did not correlate with any clinicopathological parameters. By contrast, PrPC expression was highly elevated in neoplastic compared to normal tissue and differed depending on the primary site. Interestingly, protein levels were correlated with disease recurrence (P = 0.007). Conclusively, PrPC overexpression may constitute a prognostic marker for disease recurrence and potentially a new target for anticancer therapy. However, further studies are needed to evaluate prospectively the role of PrPC expression in patients with CRC.

Acute post-stroke adiponectin in relation to stroke severity, progression and 6 month functional outcome

Abstract Background: Circulating adiponectin (ADPN) has been inversely associated with the risk of coronary artery disease and ischemic stroke (IS), due to its anti-inflammatory and anti-atherosclerotic properties. Recent experimental studies have suggested that ADPN may as well exert cerebroprotective properties in brain ischemia, therefore modifying disease outcome. We investigated whether acute post-stroke ADPN in humans might be associated with disease severity, progression and outcome. Methods: Serum ADPN was measured in 82 consecutive acute IS patients. Severity at presentation and stroke progression within the first week were evaluated according to internationally agreed definitions. Disability and functional outcomes were assessed on months 1, 3 and 6 using the modified Rankin scale (mRS) and Barthel index (BI). Additional data included information on infarct size, mortality, recurrent IS and mental state. Results: Higher ADPN was indicative of greater disability (mRS) on month 1 (OR=1·141, 95% CI=1·012–1·286, p=0·031), but this result was not replicated using the BI. ADPN was not found to be associated with either stroke severity, clinical progression, infarct size, recurrent IS, mortality, mental state, disability or functional outcome during the 6 month follow-up. Conclusions: Despite previous experimental evidence, serum ADPN measured shortly after an acute IS in humans does not seem to reliably predict disease severity, progression or outcome. The concept that circulating ADPN may beneficially modify long-term outcome of an acute IS may not be the case for human stroke.

Functional inflammatory genotypes in ischemic stroke: could we use them to predict age of onset and long-term outcome?

Functional single-nucleotide polymorphisms (SNPs) of inflammatory cytokines have been previously related to the occurrence of an ischemic stroke (IS). We investigated whether five functional SNPs (i.e., TNF-α-308G>A, IL6-174G>C, IL12B 1188A>C, IL4-589C>T, and IL10-1082G>A) might be associated with the age of onset and 6-month outcome of an acute IS. A probe-free real-time PCR methodology was used to genotype 145 consecutively admitted cases with a first-ever IS. Simple Kaplan-Mayer and adjusted Cox regression analyses showed no association between inflammatory genotypes and the age of IS onset. IL6-174G>C, IL12B 1188A>C, IL4-589C>T, and IL10-1082G>A were not found to significantly contribute to the long-term outcome of the disease. However, carriage of the TNF-α-308 GG genotype was significantly associated with reduced odds for an adverse outcome. Larger studies are needed to confirm our results.

Association of Functional VKORC1 Promoter Polymorphism with Occurrence and Clinical Aspects of Ischemic Stroke in a Greek Population

Genetic factors are considered to play an important role in determining the susceptibility to the occurrence, clinical course, and functional outcome of an acute ischemic stroke (IS). Undercarboxylation of specific vitamin K-dependent proteins, due to genetic polymorphisms of VKORC1, can affect both vascular calcification and thrombogenicity. We sought to determine the association of VKORC1 −1639G > A polymorphism with IS incidence, age of onset, severity of disease, and functional outcome after an acute IS. VKORC1 −1639G > A polymorphism was determined in 145 consecutive patients with first ever IS and 145 age- and sex-matched control subjects of Greek Caucasian origin using PCR-RFLP. Stroke severity and functional outcome were assessed on admission and at one month after stroke, respectively. Frequency of VKORC1 −1639G > A genotypes did not differ between IS patients and controls (OR = 1.12, P = 0.51). Moreover, carriage of the A allele was not associated with age of stroke onset, severity of disease (Scandinavian stroke scale score 32.2 versus 32.9, resp., P = 0.96), or poor outcome at 1 month post-stroke (52.9 versus 64.4%, resp., P = 0.31). In conclusion, VKORC1 −1639G > A polymorphism is not a genetic determinant of IS occurrence, age of onset, severity, or functional outcome of disease in a Greek population.

VEGF polymorphisms may be associated with susceptibility to colorectal cancer: A case-control study

The vascular endothelial growth factor (VEGF) has a pivotal role in angiogenesis. VEGF levels appear to be influenced by single nucleotide polymorphisms (SNPs) of the VEGF gene. The aim of this study was to assess the importance of four VEGF SNPs in modulating susceptibility to colorectal cancer. We have genotyped 223 patients with colorectal cancer and 264 healthy individuals for the -2578C>A, -1498C>T, -634G>C and +936C>T VEGF SNPs using Taqman probes in polymerase chain reactions. The -2578 A, -1498 C and -634 G alleles were more frequently detected in CRC patients compared to healthy controls. Moreover, the haplotype -2578C/-1498T was less frequent in CRC patients while the -2578A/-1498C haplotype was significantly more frequent in patients compared to healthy controls. VEGF -2578C>A and -1498C>T SNPs and -2578/-1498 haplotypes appear to be associated with susceptibility to CRC.

Association of endothelial nitric oxide synthase polymorphism G894T with functional outcome in acute stroke patients

Abstract Objectives: Endothelial nitric oxide synthase (eNOS) G894T polymorphism has been previously associated with vascular diseases including stroke, but often with conflicting results. Experimental evidence suggests that some eNOS modalities may be important in preventing stroke, while others may be important during the dynamic inflammatory processes triggered at the acute phase of stroke. Thus, we examined the possible association of eNOS G894T variation with stroke severity and functional outcome. Methods: One hundred and eight consecutive patients with first ever acute atherothrombotic or lacunar stroke, and 193 stroke-free subjects were recruited. Demographic data, medical history, and vascular risk factors were collected. Assessments for stroke severity and functional outcome were carried out on admission and at one month post-stroke, respectively. eNOS G894T genotypes were produced using a standard restriction-fragment-length polymorphism technique. Results: eNOS G894T polymorphism genotypic distributions did not differ between stroke patients and controls, as well as between each stroke subtype and controls. Carriage of T allele (GT and TT genotypes versus GG) was not significantly associated with either stroke severity or functional outcome in the univariate analysis. However, after accounting for age, gender and stroke severity, the presence of T allele strongly predicted poor outcome [odd ratio 8·49 (95% confidence intervals 1·57-46·0)]. Further adjustments for other important confounders could not alter the above significant association. Discussion: Carriers of the eNOS 894T allele are at increased risk for adverse outcome at one month post-stroke, independently of severity and other confounding factors. Therapeutic interventions targeting patients with this unfavorable mutation may be an appealing approach.