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Role of G308A polymorphism of TNF-a gene in the formation of rosacea
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Babadjanov Oyibek Abdujabbarovich, Assistant of the Department dermatology Tashkent Pediatric Medical Institute Arifov Saidkosim Saidazimovich MD, professor of dermatology head of the department of the Tashkent Institute of Advanced Medical E-mail: [email protected]
Role of G308A polymorphism of TNF-a gene in the formation of rosacea
Abstract: The results obtained by the research reliably demonstrate the presence of association of carrying rs1800629*G allele and rs1800629*G/A genotype of TNF-a gene with the risk of rosacea development. Keywords: rosacea, polymorphism rs1800629, TNF-a gene, cytokines.
Rosacea — a chronic relapsing skin disease with multifactorial nature, characterized by lesions of facial skin in the form erythema and papulo-pustular elements [8, 11]. According to the authors, the prevalence of rosacea is 10% of the world's population and holds 7th place in frequency among all dermatoses [6]. The disease mainly occurs in 30% ofpatients aged 50 years and comparably common in fair-skinned Europeans [9, 13]. In Uzbekistan, the share of rosacea among all skin diseases is 10%.
According to the classification of the National Rosacea Society (NRS) proposed in 2002 [13], there are four subtypes and one variant ofrosacea: subtype 1-erythematous-teleangiectatic rosacea; subtype 2 — papulo-pustular rosacea; subtype 3 — phymatous rosacea; subtype 4 — ocular rosacea; the variant — granulomatous rosacea.
Modern concepts of rosacea suggest that in the majority of cases the disease develops due to congenital disorders of the immune response associated with genetic defects in one or more components of the immune system [14]. This is supported by studies of some authors: 1/3 of the rosacea cases among northern Europeans is hereditary [4]. At the same time, it shows the special role of genetically mediated dysfunction of cytokine system at the core of the disease [5]. In connection with this, there has recently been notes an increased interest to the study of genetic polymorphisms ofcytokines functionally responsible for changing their protein products and the contribution ofgenotypic variants of these polymorphisms in the development and progression of rosacea.
Since the proinflammatory cytokine TNF-a plays a leading role in cellular immunity, not only promotes chemotaxis but cell proliferation, amplifying inflammation [7, 12], we considered it appropriate investigate the role of G308A polymorphism (rs1800629) of gene TNF-a in the formation of rosacea at Uzbek people. This polymorphism increases the expression of the gene and accordingly the production of the cytokine [3].
Purpose of work — study of the role of polymorphism rs1800629 of gene TNF-a in the pathogenesis of rosacea and evaluation of the prognostic value of certain genotypic variants in the development and clinical characteristics of the disease.
Methods of the research. The object of research served as the sampling of unrelated patients with "rosacea", living in different regions of the country, the total number is 140 people. The diagnosis was based on the diagnostic criteria, taking into account the major and minor signs of rosacea.
All examined patients were divided into 2 groups: I subgroup — patients with erythematous form "rosacea"; Subgroup II — patients with papulo-pustular form of "rosacea."
The control group consisted of 145 healthy unrelated individes (Uzbeks) who corresponded on the sex and age of the studied patients, and did not have in anamnesis skin disease. Genomic DNA from leucocytes of peripheral blood samples (Vacutainer Becton Dickinson International with EDTA) was isolated using the kits "QIAamp DNA Blood Mini Kit", Qiagen (Germany) and "the RNA/DNA-sorb" LLC "InterLabService" (Russia), in accordance with instruction.
Amplification of polymorphism rs1800629 of TNF-a gene was performed using thermal cyclers GeneAmp PCR-system 2720 (Applied Biosystems, USA) and Corbett Palm Cycler (Corbett Research, Australia CG1-96 model) using a commercial kit of LLC SMF Liteh (Moscow).
For genotyping of rs1800629 polymorphism of TNF-a gene specific oligonucleotide primers are used: F: 5'AATAGGTTTT-GAGGGCCATG-3'$ R: 5ATCTGGAGGAAGCGGTAGTG-3'.
Amplification was performed under the following original conditions: to pre-denature — 94 ° C (1 min. 1 cycle), 35 cycles of amplification: 94 ° C (10 sec) — denaturation, 66 ° C (20 sec) — an-
Section 6. Medical science
nealing of primers, 72 ° C (20 sec) — elongation, and final synthesis 72 ° C (1 min. 1 — cycle), storage — 10 min.
PCR products were separated by electrophoresis in 1-2% agarose gel containing ethidium bromide.
The frequency of allelic variants and genotypes (f) was calculated by the formula: f = n/2N u f = n/N, where n — the occurrence of variant (allele or genotype), N — a sample siz/Evaluation of deviation of the distributions of genotypes from the canonical distribution of Hardy-Weinberg equilibrium was performed using the computer program "GenePop".
Relative deviation the expected heterozygosity from observed (D) was calculated according to the formula: D= (h , -h )/h ,
v ' o v obs exp' exp
where h , and h — the observed and expected heterozygosity,
obs exp l J d J'
respectively.
As a tool for computing the statistical reliability of the results software package «OpenEpi 2009, Version 2.3» was used.
Results and discussion. Analysis of the polymorphism rs1800629 gene TNF-a in healthy donors Uzbeks showed no deviation of genotypes from the expected distribution at Hardy-Weinberg equilibrium (HWE) (x2=1.1, p=0.3). At the same time, the analysis of the frequency distribution of genotypes of the Hardy-Weinberg equilibrium in the group of studied patients statistically significant deviation is founded (x2=6.6; P=0.01), which is possible due to the specificity of study group (Table 1.). Deviation from the HWE can be explained by the fact that the group of patients with rosacea is not a random population sample, and selected based on the criteria of having the disease. Identify deviation, possibly due to the absence of homozygosity, ie disadvantage of homozygotes in the evaluation group at the expense of increasing the number of representatives with heterozygous variant of genotype (selective effect).
The study of differences between expected and observed frequencies of heterozygosity (D) in the studied group showed that this marker has a sufficiently high index of the heterozygous genotype. Comparative analysis of alleles and genotype frequencies of polymorphism rs1800629 gene TNF-a between the groups and subgroups of patients with rosacea, and population group were found statistically significant differences.
Frequencies of the alleles of rs1800629G and rs1800629A in the main group of patients and in the control group accounted for 82.1% and 17.8% and 92.1% and 7.9%, respectively. Here the distribution the alleles in the studied groups differed significantly, i. e. unfavorable allele rs1800629A was significantly higher in patients of the main group (x 2=12.6; P=0.0004; 0R=2.5; 95%CI 1.494-4.263) and in the subgroup of patients with erythematous stage (x2=24.7; P<0.05; OR=3.7; 95%CI 2.146-6.262).
When comparing frequencies of the alleles between patients with papulo-pustular stage ofrosacea (Il-subgroup) and the control group were not found statistically significant differences (x2=3.0; P=0.09; 0R=0.3; 95%CI 0.06867-1.29). At that sub-group I and II patients with rosacea are also very different among themselves. It was shown that rs1800629A allele frequency was significantly lower than 12.3-fold in patients with papulo-pustular stage (x2=18.1; P<0.05; 0R=12.32; 95%CI 2.916-52.01).
When comparing samples of patients and appropriate control are also found statistically significant differences in the distribution of genotype frequencies of this locus. In the group of patients the frequency of adverse heterozygous genotype rs1800629*G/A gene TNF-a was significantly higher compared to the control (35.7%
versus 15.9%, x2=14.3; P<0.05; 0R=3.0; 95%CI 1.677-5.179). According to the calculated odds ratio, risk of development of rosacea in carriers of this genotype in 3.0 times higher than in carriers of the homozygous wild genotype.
When compared with the subgroup of patients with erythem-atous stage with population sample also are revealed statistically significant differences (p <0.05). The frequency of heterozygous genotype rs1800629*G/A in these groups was 48.0% and 15.9%, respectively. In analyzing the distribution of the genotype frequencies in group II patients and the control group showed no statistically significant differences (x2=3.2; P=0.07; 0R=0.3; 95%CI 0.06292-1.239).
It should be noted that the frequency of wild rs1800629*G/G genotype in the main group and subgroup I patients with rosacea was significantly higher than in controls (P <0.05), which may be associated with a decreased risk of developing the disease in carriers of this genotype. Here founded some decrease the frequency of a given genotype in subgroup II compared to the control group (84.1% versus 95.0%, respectively), but the differences were not statistically significant (x2=3.2; P=0.07; 0R=3.5; 95%CI 0.8073-15.89).
Comparative analysis of the genotypes frequencies of polymorphism rs1800629 between subgroups of patients with rosacea showed statistically significant differences (p<0.05). The frequencies of homo- and heterozygous genotypes in the subgroups of patients studied were: 52.0%, 48.0% and 0.0% — in subgroup I, 95.0%, 5.0% and 0.0% — in subgroup II (x2=23.1; P<0.05; 0R=17.5; 95%CI 4.013-76.66). In the studied groups of patients and control homozygous genotype rs1800629*A/A of gene TNF-a was not identified. Thus, received during of the study results reliably indicate the presence of the association carrier state of allele rs1800629*G and genotype rs1800629*G/A of TNF-a gene with the risk of rosacea.
Conclusion. Rs1800629 polymorphism of of TNF-a gene in patients with rosacea and conditionally healthy donors Uzbeks, characterized by wide allelic diversity. Functionally unfavorable genotype rs1800629*G/A of TNF-a gene significantly contributes to the formation of rosacea and more associated with erythematous stage of rosacea. The risk of development of pathology in the presence of this genotype may be increased more than 3 times, and the chances of forming erythematous stage of disease when it present are increased by more than 4.5 times (x2=29.7; P<0.05; 0R=4.9; 95%CI 2.704-8.865). At that, wild genotype rs1800629*G/G of TNF-a gene may be associated with a decreased risk of developing the disease in carriers of this genotype.
It should, however, admit that today pathogenesis based concept not only for of rosacea, but most other dermatoses, missing. In the literature specific work on the study of polymorphism rs1800629 of TNF-a gene in patients with rosacea are virtually absent. The number of papers devoted to assessing the role cytokines genes in the formation of other dermatoses, few, and their results are controversial [1, 2. 10].
However, despite a certain fragmentation, similar studies in the future open new horizons in identifying etiopathogenesis of dermatoses, including rosacea. We consider it necessary to conduct further studies of other cytokines genes, which would bind together genetic and other mechanisms for the development rosacea and to develop not only effective means of forecasting the development and heaviness of diseases of the autoimmune nature, but also to identify perspective methods of their personalized therapy.
Physical fitness of preschool educational institutions children of Uzbekistan
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2. Al-Shobaili HA, Salem TA, Alzolibani AA, Robaee AA, Settin AA. Tumor necrosis factor-alpha -308 G/A and interleukin 10 -1082 A/G gene polymorphisms in patients with acne vulgaris. J Dermatol Sci 2012;68:52-5.
3. Bayley JP, Ottenhoff TH, Verweij CL Is there a future for TNF promoter polymorphisms? Genes Immun 2004;55:315-29.
4. Del Rosso JQ. Update on rosacea pathogenesis and correlation with medical therapeutic agents. Cutis 2006;78:97-00.
5. Gerber PA, Buhren BA, Steinhoff M, Homey B: Rosacea: the cytokine and chemokine network. J Investig Dermatol Symp Proc. 2011;15:40-7.
6. Gupta AK, Chaudhry MM Rosacea and its management: an overview. J Eur Acad Dermatol Venereol 2005;19:273-85.
7. Gupta S, Gollapudi S. Molecular mechanisms of TNF-alpha-induced apoptosis in aging human T cell subsets. Int J Biochem Cell Biol. 2005;37 (5):1034-42.
8. Korting HC, Schollmann C. Current topical and systemic approaches to treatment of rosacea. J Eur Acad Dermatol Venereol. 2009a;23:876-82.
9. Powell FC. Clinical practice. Rosacea. N Engl J Med. 2005;352:793-03.
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11. van Zuuren EJ, Kramer S, Carter B, Graber MA, Fedorowicz Z. Interventions for rosacea. Cochrane Database Syst Rev 2011;3: CD003262.
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13. Wilkin J, Dahl M, Detmar M, Drake L, Feinstein A, Odom R, et al. Standard classification of rosacea: Report of the National Rosacea Society Expert Committee on the Classification and Staging of Rosacea. J Am Acad Dermatol. 2002;46:584-7.
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Basharova Laylo Maratovna, Research institute of sanitation, hygiene and occupational diseases of Ministry of Health of the Republic of Uzbekistan, Tashkent, Uzbekistan
E-mail: [email protected]
Physical fitness of preschool educational institutions children of Uzbekistan
Abstract: 3112 children at the age of 5 and 6 years which are brought up in preschool educational institutions of Tashkent city, 3 areas of the republic (Tashkent, Navoy, Surkhandarya) and Karakalpakstan have been examined. Indicators of physical readiness of children by results of 8 control tests have been studied. The average rates testing results are depended on age, sex and the residence region: at children from 5 to 6 years improvement of physical fitness was observed; the number of boys who could not execute test physical exercises, was almost in 10 times more, than their contemporaries; the highest rates of physical fitness were observed among preschool children of Tashkent city, the lowest — among children of Karakalpakstan. Keywords: children from 3 to 6 years preschool educational institutions, physical fitness.
In the Uzbekistan Republic is worked specially developed training programs on physical training intended for children of different age groups. Results of scientific works of the republic scientists are evidenced by some types of exercises the control standards for physical training expounding in the training program are exceed the level of the actual average indicators of physical fitness of the children who are engaged in the main group of physical training [1, 26]. Organized and free motion activity are effected on development and improvement of physical qualities, improvement of the children's health, including enhancement cardiovascular, respiratory and nervous systems of activity, strengthening of the locomotive system, improving of a metabolism, resistance upgrading to diseases and mobilization of protective forces of an organism [3, 51-54; 4, 51-53; 5, 4-9; 6, 2492-2495].
Goal: to reveal features of physical fitness of 5 and 6 years old children, living in the Uzbekistan and scientifically substantiate the
efficiency of implementation developed complex of physical exercises.
Materials and methods researches. It has been surveyed children who are brought up in preschool educational institutions of Tashkent city, Tashkent, Navoiy and Surkhandarya areas of the republic and Karakalpakstan. In total were examined 3112 children, including 1651 boys and 1461 girls. The amount of 5 and 6 years children were almost the same.
Physical fitness level of children has been estimated by 8th control tests results which have been included such exercises as: lifting ofdirect close feet to vertical level from the state "lying on a back, hands behind the head" and releasing them to a floor (times); skipping rope in 30 seconds (times); hoop rotation round a waist in one minute (time); a throw of a tennis ball (5 efforts) in the purpose (basket) located at 6 m distance from the child (times); long jumps from a place (cm); jogging on 30 meters distance (s); chinning up in hanging on high (boys) and low (girl) crossbeams (times); bouncing on a foot in 30 seconds (times).
