Levels of some cytokines in subretinal fluid in proliferative vitreoretinopathy and rhegmatogenous retinal detachment.

PURPOSE: To determine the cytokine-mediated pathways of inflammation in the clinical course of proliferative vitreoretinopathy (PVR). METHODS: Cytokines IL-2, IL-6, INF-gamma were measured by enzyme-linked immunosorbent assays. Aspirates of subretinal fluid were obtained from 14 eyes with PVR and six with uncomplicated rhegmatogenous retinal detachments. Aspirates obtained via pars plana from ten cadaver vitreouses of normal eyes were used as controls. RESULTS: Subretinal fluids from eyes with PVR, uncomplicated rhegmatogenous retinal detachment and cadaver vitreous contained low concentrations of IL-2, with inconsequential differences between groups. IL-6 and IFN-gamma were twice as high in the subretinal fluid from eyes with rhetmatogenous retinal detachment as in cadaver vitreous. INF-gamma was elevated up to six times in eyes with PVR compared to controls. Fibronectin was found in a larger proportion of eyes with PVR than with uncomplicated rhegmatogenous retinal detachment and cadaver vitreous from healthy persons. CONCLUSIONS: These are only preliminary investigations and are too few for statistical analysis. The results suggest that cytokine-mediated pathways of inflammation are involved in the pathogenesis of rhegmatogenous retinal detachment and in cellular interactions leading to the development of PVR.

[Localization of conserved and variable segments in amino acid sequences of homologous proteins using a personal computer]. / Lokalizatsiia konservativnykh i variabel'nykh uchastkov v aminokislotnykh posledovatel'nostiakh gomologichenykh belkov na personal'nom komp'iutere.

A set of aligned homologous protein sequences is divided into two groups consisting of the most related sequences m and k. The value of the position variability of homologous protein sequences is defined as a number of failures to coincide in the intergroup comparison of all possible k x m pairs of amino acid residues in that position divided by k x m. The position variability value plotted vs the sequence position number with a window of 10 positions gives the intergroup local variability profile. The area S of the figure included between the local variability profile and the straight line corresponding to the mean local variability value is compared with the average area S(r) for 1000 random homologous protein families. If S is greater than S(r) by more than 2 standard deviation units sigma r the local variability profile is assumed to contain peaks and hollows corresponding to significant variable and conservative regions of the sequences. The profile extrema containing the area surplus delta S = S-(S(r) + 2 sigma r) are cut off by two straight lines to locate significant regions. The numerical experiment on the family of homologous phospholipases A2 revealed the linear dependence of the values S(r) and sigma r upon the position variability standard deviation sigma v of the homologous sequences. Furthermore, it was shown for protein families of various length (rhodopsins, aspartate aminotransferases, cytochromes b, L- and M-subunits of photosynthetic bacteria photoreaction centre and alpha-subunits of Na, K-ATPase), that delta S = S - n(S'r + 2 sigma r), where S - the area of the local variability profile, n = L/l (L - the length of the given protein family and l - the length of the hypothetical protein domain). If l = 250 then S'r = -1.42 + 62.56 sigma v and sigma'r = -0.14 + 7.46 sigma v.

The significant conservative and variable regions of the homologous protein sequences.

A method of identification of significant conservative and variable regions in homologous protein sequences is presented. A set of aligned homologous sequences is divided into two groups consisting of m and n most related sequences. Each pair of sequences from different group is compared using unitary similarity matrix. The superposition of pairwise comparisons scanned by a window of 10 amino acid residues gives intergroup local variability profile (VP). Area S of the figure between the VP and its mean value line is compared with averaged area S(r) of 1000 VPs of artificial homologous protein families. The difference (S-S(r)) given in standard deviation units sigma r is believed to be the amino acid substitution overall irregularity along the homologous protein sequences OI = (S-S(r))/sigma r. If OI greater than 2, the real VP extrema containing the surplus of area S-(S(r) + 2 sigma r) are cut off. The cut off stretches are likely to be significant conservative and variable regions. The significant conservative and variable regions of six homologous sequence families (phospholipases A2, cytochromes b, alpha-subunits of Na, K-ATPase, L- and M-subunits of photosynthetic bacteria photoreaction centre and human rhodopsins) were identified. It was shown that for artificial homologous protein sequences derived by k-fold lengthening of natural proteins the OI value rises as square root of k. To compare the degree of substitution irregularity in homologous protein sequence families of different length L the value of standard substitution overall irregularity for L = 250 is proposed.

Conservative and variable regions of homologous snake phospholipases A2 sequences: prediction of the taxon-specific peptides structure.

Homologous amino acid sequences of phospholipases A2 (PLA2) of snakes belonging to the families Elapidae, Viperidae, and Colubridae were considered in order to study the conservative and variable regions location. The PLA2 sequences were divided into two groups (taxons) according to the phylogenetic tree reconstructed from the pair similarity matrix. Results of the intergroup comparison were plotted to facilitate the identification of significant conservative and variable regions. It was shown that the results of the comparison between two phylogenetic groups of snake PLA2 did not much depend on the number of each group representatives and did not markedly change if one of the groups was represented by the single sequence. The knowledge of the number and location of conservative and variable regions and their dependence on the phylogenetic relations between compared taxa may be used to predict a synthetic peptide structure to obtain specific antibodies against PLA2 of one of these taxons. Such prediction is possible if there is a specific region conservative for one taxon but variable for two of them.

[Conserved and variable segments of the snake phospholipase A2 amino acid domain]. / Konservativnye i variabel'nye uchastki aminokislotnykh posledovatel'nostei fosfolipaz A2 zmei.

Homologous amino acid sequences of phospholipases A2 of snakes belonging to families Elapidae, Viperidae and Colubridae were considered in order to study the location of conservative and variable regions. To identify significant conservative and variable regions a comparison between two groups of aligned sequences of snake phospholipases A2 was successfully applied. The phospholipases A2 sequences were divided into two groups (taxons) according to the phylogenetic tree reconstructed from the pair distance matrix. Results of the comparison were plotted to facilitate the identification of significant conservative and variable regions. It was shown, that the results of the comparison between two phylogenetic groups of snake phospholipases A2 didn't depend much on the number of each group representatives, and the location of conservative and variable regions didn't significantly change if one of the groups was represented by the single sequence. It should be mentioned, that the more the phylogenetic difference between groups of phospholipases A2 the more was the number of significant conservative and variable regions. The knowledge of the number and location of conservative and variable regions and their dependence on phylogenetic relations between the compared taxons can be used to predict the synthetic peptide structure to obtain antibodies of various specificity. These antibodies may have either a wide range of cross-reactivity against all of phospholipases A2 or a limited range of cross-reactivity against phospholipases A2 of one taxon.

[A method of detecting conserved and variable segments in the amino acid sequences of homologous proteins]. / Metod obnaruzheniia konservativnykh i variabel'nykh uchastkov v aminokislotnykh posledovatel'nostiakh gomologichnykh belkov.

A set of aligned homologous protein sequences is divided into two groups consisting of m and n sequences. Each group contains sequences from the most related organisms. Value of the position dissimilarity of proteins from different groups of m and n sequences is defined as a number of mismatches in comparison of all possible m X n pairs of amino acid residues in the position (each from different group) divided by m X n. Ten position average of dissimilarity values is plotted vs. the first position number. Area of the figure between the profile of dissimilarity values and its mean value line characterizes the overall irregularity of amino acid substitutions along the protein sequences. If the area is greater than the average area for 1000 random profiles by more than two standard deviation units, the profile extrema containing the "surplus" of area are cut off. The cut-off stretches are likely to be variable and constant regions. If necessary, each of stretches may be separately tested and statistically estimated using a standard size sample of artificial protein families. Intergroup comparison of protein sequences reveals high overall irregularity of amino acid substitutions and identifies variable and conservative regions for all considered families of proteins: phospholipases A2, aspartate aminotransferases, alpha-subunits of Na+, K(+)-ATPase, L- and M-subunits of photosynthetic bacteria photoreaction centre, human rhodopsins.

Identification of significant conservative and variable regions in homologous protein sequences.

A set of aligned homologous protein sequences is divided into 2 groups consisting of m and n sequences. Each group contains sequences from the most related species. Value of the position variability of homologous proteins sequences is defined as a number of failures to coincide in comparison of all possible m.n pairs of amino acid residues (each from a different group) in that position divided by m.n. The variability value plotted vs the sequence position number with a window of 10 positions gives the intergroup variability profile (VP). Area of the figure included between the VP and its mean value line characterizes the overall irregularity of amino acid substitutions along the protein sequences. If the area value S is greater than the average area Sr for 1000 random VPs by more than 2 standard deviation units (sigma), the real VP extrema containing the surplus of area S-(Sr + 2 sigma) are cut off. The cut-off stretches are likely to be significant variable and conservative regions. Intergroup comparisons of protein sequences reveal high overall irregularity of amino acid substitutions and identify variable and conservative regions for all considered families of proteins: phospholipases A2, aspartate aminotransferases, alpha-subunits of Na+,K(+)-ATPase, L- and M-subunits of photosynthetic bacteria photoreaction centre, and human rhodopsins.

[Graphic identification of conservative and variable segments in the amino acid sequences of homologous proteins]. / Graficheskaia identifikatsiia konservativnykh i variabel'nykh uchastkov v aminokislotnykh posledovatel'nostiakh gomologichnykh belkov.

An algorithm is presented for localizing variable and constant regions in homologous protein sequences. A set of aligned protein sequences is divided into two groups consisting of m and n sequences. Each group contains sequences of most related species. Value of the position dissimilarity of proteins from different groups of m and n sequences is defined as a number of failures to coincide in comparison with all possible mXn pairs of amino acid residues in the position (each from different group) divided by mXn. The position dissimilarity value of m protein sequences within a group is defined as the number of failures to coincide in comparison with all possible mX X(m-1)/2 pairs of amino acid residues divided by mX(m-1)/2. Ten position average of dissimilarity values is plotted vs. the first position number. Area of the figure included between the profile of dissimilarity values and its mean value line characterizes the overall irregularity of amino acid substitutions along the protein sequences. If the area value is greater than the average area for 1000 random profile by more than two standard deviation units, the profile extrema containing the "surplus" of area are cut off. The cut off stretches are likely to be variable and constant regions. In case of "between groups" comparisons it is found that the overall irregularity of amino acid substitutions is very high for all considered families of proteins; phospholipases A2, aspartate aminotransferases, alpha-subunits of Na+,K(+)-ATPase, L- and M-subunits of photosynthetic bacteria photoreaction centre, human rhodopsins.