Polymorphisms in some proinflammatory genes (TNFα and ß, IL-1ß, IL-6, ADAM17) in severe chronic venous disease.

BACKGROUND: Chronic venous disease (CVD) is a common disorder of lower extremities. OBJECTIVES: The study was scheduled to investigate the relationship between polymorphisms in major proinflammatory genes TNF α (-238 A/G; -308 A/G), TNF ß (NcoI), IL-1ß (+3953 T/C); IL-6 (-174 G/C; -596 G/C) and ADAM17 (3'TACE) and CVD risk. Genotype-phenotype study was calculated to test possible association between examined genotypes and phenotypes of CVD. METHODS: Finally, 150 CVD patients and 227 control subjects were enrolled to the study. Genotypes in proinflammatory gene polymorphisms were identified from isolated DNA by PCR method and restriction analysis. RESULTS: Significant differences in genotype distribution/allelic frequencies in TNF ß gene, IL-1 ß gene and in ADAM17 gene polymorphisms were found between CVD women and control ones. In the genotype-phenotype study, identified genotypes were associated with arterial hypertension (ADAM17, IL-6-men), ischaemic heart disease (TNF α and ß genes), diabetes mellitus (ADAM17-women, TNF ß-men), age of CVD onset (TNF α and IL-6), ulceration (ADAM17), duration of ulceration (ADAM17), ulceration recurrence (ADAM17-women), home care necessity (TNF α), varices surgery (TNF α), erysipelas development (ADAM17-men) and tumour development (TNF α). CONCLUSION: Studying of these polymorphisms associations can help us better identify patients at higher risk of developing severe CVD.

Hidden Potential of Highly Efficient and Widely Accessible Thrombolytic Staphylokinase.

Stroke burden is substantially increasing but current therapeutic drugs are still far from ideal. Here we highlight the vast potential of staphylokinase as an efficient, fibrin-selective, inexpensive, and evolvable thrombolytic agent. The emphasis is escalated by new recent findings. Staphylokinase nonimmunogenic variant was proven noninferior to alteplase in a clinical trial, with decreased risk of intracranial hemorrhage and the advantage of single bolus administration. Furthermore, our detailed kinetic analysis revealed a new staphylokinase limiting bottleneck whose elimination might provide up to 1000-fold higher activity than the clinically approved alteplase. This knowledge of limitations unlocks new possibilities for improvements that are now achievable by the community of protein engineers who have the required expertise and are ready to transform staphylokinase into a powerful molecule. Collectively, the noninferiority and safety of nonimmunogenic staphylokinase together with the newly identified effectivity limitation make staphylokinase a perfect candidate for further exploration, modification, and advancement to make it the next-generation widely accessible thrombolytic drug effectively treating stroke all around the world, including middle- and low-income countries.

Identification, characterization, and engineering of glycosylation in thrombolytics.

Cardiovascular diseases, such as myocardial infarction, ischemic stroke, and pulmonary embolism, are the most common causes of disability and death worldwide. Blood clot hydrolysis by thrombolytic enzymes and thrombectomy are key clinical interventions. The most widely used thrombolytic enzyme is alteplase, which has been used in clinical practice since 1986. Another clinically used thrombolytic protein is tenecteplase, which has modified epitopes and engineered glycosylation sites, suggesting that carbohydrate modification in thrombolytic enzymes is a viable strategy for their improvement. This comprehensive review summarizes current knowledge on computational and experimental identification of glycosylation sites and glycan identity, together with methods used for their reengineering. Practical examples from previous studies focus on modification of glycosylations in thrombolytics, e.g., alteplase, tenecteplase, reteplase, urokinase, saruplase, and desmoteplase. Collected clinical data on these glycoproteins demonstrate the great potential of this engineering strategy. Outstanding combinatorics originating from multiple glycosylation sites and the vast variety of covalently attached glycan species can be addressed by directed evolution or rational design. Directed evolution pipelines would benefit from more efficient cell-free expression and high-throughput screening assays, while rational design must employ structure prediction by machine learning and in silico characterization by supercomputing. Perspectives on challenges and opportunities for improvement of thrombolytic enzymes by engineering and evolution of protein glycosylation are provided.

Matrix metalloproteinase-2 promoter variability in psoriasis.

The expression of matrix metalloproteinase-2 was observed to be significantly upregulated in psoriasis. The aim of this study was to associate the DNA polymorphic variants in MMP-2 promoter gene with psoriasis and/or with psoriasis phenotypes related to psoriasis and comorbid heredity. In the total of 582 Czech Caucasian individuals (386 patients with psoriasis and 196 controls of similar age and sex distribution without personal or family history of chronic disease of the skin), four MMP-2 promoter polymorphisms (-1575G/A, -1306C/T, -790T/G and -735C/T) were detected by PCR methods. A significant association of GG genotype of -790 MMP-2 polymorphism with psoriasis was observed (Pcorr = 0.04). Although no significant case-control differences in frequency of associated GG(-1575)CC(-1306)TT(-790) MMP-2 promoter genotype were observed, the genotype was found to be significantly less frequent in patients with family history of psoriasis (close as well as distant), family history of diabetes and personal history of allergy (2/11 vs. 55/32, odds ratio (OR) for GGCCTT 0.11, 95% confidential interval 0.02-0.50, Pcorr = 0.01). The significant difference between psoriatic patients with positive anamnestic data on diabetes, psoriasis and allergy compared with psoriatic patients that have only positive family history of diabetes was also observed (2/11 vs. 38/31, P = 0.009, Pcorr = 0.04; OR 0.15, 95% CI = 0.03-0.72 for psoriatic patients with GGCCTT genotype and family history of psoriasis, diabetes and personal history of allergy). To conclude, the associated GGCCTT genotype in the promoter of MMP-2 gene was less frequent in patients with positive family history of psoriasis, diabetes and personal history of allergy compared with psoriatic patients without them (2/11 vs. 68/57, P = 0.007, Pcorr = 0.04; OR = 0.15, 95% CI = 0.03-0.72 for psoriatic patients with family history of psoriasis and diabetes and with allergy). Based on our results, we suggest that the MMP-2 located in the psoriasis susceptibility region on 16q (psoriasis susceptibility 8, PSORS8) should be considered as a gene modulator of psoriasis in specific subgroups of patients. In the future, similar genetic characteristics could contribute to the data assembly of genetic predisposition to psoriasis and could lead to therapy improvement based on time-proved individual pharmacogenetic aspects detected in psoriasis patients.