Aquo-pentamine Co(III) complexes as models for carbonic anhydrase

The hydrolysis of 4-nitrophenyl acetate by metal complexes Co(en)2(imH)H2O3+, Co(en)2(bzmH)H2O3+, and Co(en)2(imCH3)H2O3+ (imH = imidazole, bzmH = benzimodazole, imCH3 = methyl imidazole) has been investigated in the pH range 5.4-8.9. The small difference in nucleophilic reactivity in the pH range 5.4-6.7 is assumed to be due to hydrogen bonding abilities of the imidazole and substituted imidazole ligands and small pKa differences (k2(imH) = 2.2 X 10(-2) M-1 sec-1, k2(bzmH) = 5.68 X 10(-2) M-1 sec-1, k2(imCH3) = 1.35 X 10(-2) M-1 sec-1, 40 degrees C, 1 = 0.3 NaClO4, pKa(imH) = 6.2, pKa(imCH3) = 6.2 and pKa(bzmH) = 5.9). In the pH range 7.8-8.9, the differences in nucleophilic reactivity (k3(imH) = 85.5 X 10(-2) M-1 sec-1, k3(bzmH) = 33.4 X 10(-2) M-1 sec-1, 40 degrees C, I = 0.3 NaClO4) are reconciled with a significant steric factor outweighing the acidity of the benzimidazole complex. In the pH region 6.7-7.7, the deviation from linearity is presumably due to both hydroxo and imido ligands functioning as nucleophiles, the latter being about 40 times stronger than the former.     

Biogenic and synthetic polyamines bind cationic dendrimers

Biogenic polyamines are essential for cell growth and differentiation, while polyamine analogues exert antitumor activity in multiple experimental model systems, including breast and lung cancer. Dendrimers are widely used for drug delivery in vitro and in vivo. We report the bindings of biogenic polyamines, spermine (spm), and spermidine (spmd), and their synthetic analogues, 3,7,11,15-tetrazaheptadecane.4HCl (BE-333) and 3,7,11,15,19-pentazahenicosane.5HCl (BE-3333) to dendrimers of different compositions, mPEG-PAMAM (G3), mPEG-PAMAM (G4) and PAMAM (G4). FTIR and UV-visible spectroscopic methods as well as molecular modeling were used to analyze polyamine binding mode, the binding constant and the effects of polyamine complexation on dendrimer stability and conformation. Structural analysis showed that polyamines bound dendrimers through both hydrophobic and hydrophilic contacts with overall binding constants of K(spm-mPEG-G3) = 7.6 × 10(4) M(-1), K(spm-mPEG-PAMAM-G4) = 4.6 × 10(4) M(-1), K(spm-PAMAM-G4) = 6.6 × 10(4) M(-1), K(spmd-mPEG-G3) = 1.0 × 10(5) M(-1), K(spmd-mPEG-PAMAM-G4) = 5.5 × 10(4) M(-1), K(spmd-PAMAM-G4) = 9.2 × 10(4) M(-1), K(BE-333-mPEG-G3) = 4.2 × 10(4) M(-1), K(Be-333-mPEG-PAMAM-G4) = 3.2 × 10(4) M(-1), K(BE-333-PAMAM-G4) = 3.6 × 10(4) M(-1), K(BE-3333-mPEG-G3) = 2.2 × 10(4) M(-1), K(Be-3333-mPEG-PAMAM-G4) = 2.4 × 10(4) M(-1), K(BE-3333-PAMAM-G4) = 2.3 × 10(4) M(-1). Biogenic polyamines showed stronger affinity toward dendrimers than those of synthetic polyamines, while weaker interaction was observed as polyamine cationic charges increased. The free binding energies calculated from docking studies were: -3.2 (spermine), -3.5 (spermidine) and -3.03 (BE-3333) kcal/mol, with the following order of binding affinity: spermidine-PAMAM-G-4>spermine-PAMMAM-G4>BE-3333-PAMAM-G4 consistent with spectroscopic data. Our results suggest that dendrimers can act as carrier vehicles for delivering antitumor polyamine analogues to target tissues.     

Thyroxine-protein interactions. Interaction of thyroxine and triiodothyronine with human thyroxine-binding globulin.

The effect of temperature on the binding of thyroxine and triiodothyronine to thyroxine-binding globulin has been studied by equilibrium dialysis. Inclusion of ovalbumin in the dialysis mixture stabilized thyroxine-binding globulin against losses in binding activity which had been found to occur during equilibrium dialysis. Ovalbumin by itself bound the thyroid hormones very weakly and its binding could be neglected when analyzing the experimental results. At pH 7.4 and 37 degrees in 0.06 M potassium phosphate/0.7 mM EDTA buffer, thyroxine was bound to thyroxine-binding globulin at a single binding site with apparent association constants: at 5 degrees, K = 4.73 +/- 0.38 X 10(10) M-1; at 25 degrees, K = 1.55 +/- 0.17 X 10(10) M-1; and at 37 degrees, K = 9.08 +/- 0.62 X 10(9) M-1. Scatchard plots of the binding data for triiodothyronine indicated that the binding of this compound to thyroxine-binding globulin was more complex than that found for thyroxine. The data for triiodothyronine binding could be fitted by asuming the existence of two different classes of binding sites. At 5 degrees and pH 7.4 nonlinear regression analysis of the data yielded the values n1 = 1.04 +/- 0.10, K1 = 3.35 +/- 0.63 X 10(9) M-1 and n2 = 1.40 +/- 0.08, K2 = 0.69 +/- 0.20 X 10(8) M-1. At 25 degrees, the values for the binding constants were n1 = 1.04 +/- 0.38, K1 = 6.5 +/- 2.8 X 10(8) M-1 and n2 = 0.77 +/- 0.22, K2 = 0.43 +/- 0.62 X 10(8) M-1. At 37 degrees where less curvature was observed, the estimated binding constants were n1 = 1.02 +/- 0.06, K1 = 4.32 +/- 0.59 X 10(8) M-1 and n2K2 = 0.056 +/- 0.012 X 10(8) M-1. When n1 was fixed at 1, the resulting values obtained for the other three binding constants were at 25 degrees, K1 = 6.12 +/- 0.35 X 10(8) M-1, n2 = 0.72 +/- 0.18, K2 = 0.73 +/- 0.36 X 10(8) M-1; and at 37 degrees K1 = 3.80 +/- 0.22 X 10(8) M-1, n2 = 0.44 +/- 0.22, and K2 = 0.43 +/- 0.38 X 10(8) M-1. The thermodynamic values for thyroxine binding to thyroxine-binding globulin at 37 degrees and pH 7.4 were deltaG0 = -14.1 kcal/mole, deltaH0 = -8.96 kcal/mole, and deltaS0 = +16.7 cal degree-1 mole-1. For triiodothyronine at 37 degrees, the thermodynamic values for binding at the primary binding site were deltaG0 = -12.3 kcal/mole, deltaH0 = -11.9 kcal/mole, and deltaS0 = +1.4 cal degree-1 mole-1. Measurement of the pH dependence of binding indicated that both thyroxine and triiodothyronine were bound maximally in the region of physiological pH, pH 6.8 to 7.7.     

Steady-state properties of calcium binding to parvalbumins from bullfrog skeletal muscle: effects of Mg2+, pH, ionic strength, and temperature

To improve our understanding of the physiological roles of parvalbumins, PA-1 (pI 4.78) and PA-2 (pI 4.97) parvalbumins were prepared from bullfrog skeletal muscle and their calcium binding properties were examined in a medium of constant ionic strength (I = 0.106, pH 6.80, at 20 degrees C) containing various concentrations of Mg2+ by using a metallo-indicator, tetramethylmurexide. Apparent binding constants for Ca2+ in the presence of Mg2+ changed in the manner expected if Ca2+ and Mg2+ compete for two independent homogeneous binding sites. The following values were obtained: for PA-1, KCa = 1 X 10(7) M-1, KMg = 900 M-1; for PA-2, KCa = 6 X 10(6) M-1, KMg = 830 M-1 (I = 0.106, pH 6.80, at 20 degrees C). The apparent binding constants are strongly dependent on temperature: at 10 degrees C for PA-1, KCa = 2 X 10(8) M-1, KMg = 10(4) M-1; for PA-2, KCa = 5 X 10(7) M-1, KMg = 5 X 10(3) M-1 (I = 0.106, pH 6.80). The dependence of the affinities for Ca2+ on ionic strength is similar to or less than that of GEDTA (EGTA). The affinities for Ca2+ and Mg2+ of parvalbumins are unchanged between pH 6.5 and 7.2.     

Steady-state kinetics of thiocyanate oxidation catalyzed by human salivary peroxidase

A steady-state kinetic analysis was made of thiocyanate (SCN-) oxidation catalyzed by human peroxidase (SPO) isolated from parotid saliva. For comparative purposes, bovine lactoperoxidase (LPO) was also studied. Both enzymes followed the classical Theorell-Chance mechanism under the initial conditions [H2O2] less than 0.2mM, [SCN-] less than 10mM, and pH greater than 6.0. The pH-independent rate constants (k1) for the formation of compound I were estimated to be 8 X 10(6) M-1 s-1 (SD = 1, n = 18) for LPO and 5 X 10(6) M-1 s-1 (SD = 1, n = 11) for SPO. The pH-independent second-order rate constants (k4) for the oxidation of thiocyanate by compound I were estimated to be 5 X 10(6) M-1 s-1 (SD = 1, n = 18) for LPO and 9 X 10(6) M-1 s-1 (SD = 2, n = 11) for SPO. Both enzymes were inhibited by SCN- at pH less than 6. The pH-independent equilibrium constant (Ki) for the formation of the inhibited enzyme-SCN- complex was estimated to be 24 M-1 (SD = 12, n = 8) for LPO and 44 M-1 (SD = 4, n = 10) for SPO. An apparent pH dependence of the estimated values for k4 and Ki for both LPO and SPO was consistent with a mechanism based on assumptions that protonation of compound I was necessary for the SCN- peroxidation step, that a second protonation of compound I gave an inactive form, and that the inhibited enzyme-SCN- complex could be further protonated to give another inactive form.(ABSTRACT TRUNCATED AT 250 WORDS)     

Vitamin A and glutathione-mediated free radical damage: competing reactions with polyunsaturated fatty acids and vitamin C

Vitamin A (retinol reacts extremely rapidly (k = 1.4 x 10(9) M-1 s-1) with thiyl free radicals derived from glutathione to form a free radical with a very strong visible absorption (lambda max. = 380 nm, E max. = 4.0 x 10(4) M-1 cm-1). Arachidonate, linolenate, linoleate and ascorbate also react readily but much more slowly (k = 2.2 x 10(7), 1.9 x 10(7), 1.3 x 10(7) and 3.6 x 10(8) M-1 s-1 respectively). These results support the possibility that vitamin A might play a role in protecting lipid membranes against thiyl free radical mediated damage.     

锦鸡儿属植物14个种类的核型

报道了锦鸡儿属Caragana 14种的核型,体细胞中期染色体的核型分别为：（1）柠条Caragana korshinskii Kom.2n=16=19m 7sm;(2)印度锦鸡儿C.gerardiana Royle 2n=16=12m 4sm;(3）锦鸡儿C.sinica(Buc‘hoz)Rehd。2n=24=15m 9sm;(4)狭叶锦鸡儿C.stenophylla Pojark 2n=32=21m 10sm 1st;(5)短脚锦鸡儿C.brachypoda Pojark.2n=16=8m 5sm 3st;(6)云南锦鸡儿C.franchetiana Kom.2n=16=13m 2sm 1st;(7)甘蒙锦鸡儿C.opulens Kom.2n=16 1b=8m 7sm 1st 1B;(8)刺叶锦鸡儿C.acanthopylla Pojark.2n=16=12m 4sm;(9)红花锦鸡儿C.rosea Maxim.2n=16=10m 6sm;(10)扁刺锦鸡儿C.boisi Schneid.2n=16=11m 4sm 1st;(11)二色锦鸡儿C.bicolor Kom.2n=16 1b=10m 6sm 1B;（12）川西锦鸡儿C.erinacea Kom.2n=16=12m 3sm 1st;（13）粗刺锦鸡儿C.crassispina Marq.2n=16=10m 6sm;(14)树锦鸡儿C.arborescens Lam.2n=16 1b=10m 6sm 1b.其中短脚锦鸡儿、云南锦鸡儿、红花锦鸡儿、扁刺锦鸡儿、二色锦鸡儿、粗刺锦鸡儿的核型为首次报道。     

Interaction of metal ions with caffeine and theophylline: stability and structural features

The interactions of caffeine and theophylline with divalent cadmium, mercury, strontium and barium ions were studied in aqueous solution and physiological pH. Fourier transform infrared spectroscopy (FTIR) and absorption spectra were used to determine the cation binding mode and association constants. Spectroscopic results showed that Cd(2+), Hg(2+), Sr(2+) and Ba(2+) bind strongly to caffeine and theophylline. Direct and indirect (through metal hydration shell) interactions were observed for caffeine and theophylline with Cd(2+), Hg(2+), Sr(2+) and Ba(2+) through O6 and N9 (caffeine) and O6, N9 and N7 atoms (theophylline). The overall binding constants are:k(Cd-caffeine) = 1.24 x 10(5) M(-1), k(Hg-caffeine) = 1.74 x 10(5) M(-1), k(Sr- caffeine) = 3.3 x 10(4) M(-1), k(Ba-caffeine) = 1.8 x 10(4) M(-1), k(Cd-theophylline) = 5.75 x 10(5) M(-1), k(Hg-theophylline) = 2.14 x 10(5) M(-1), k(Sr-theophylline) = 4.6 x 10(4) M(-1), k(Ba-theophylline) = 3 x 10(4) M(-1). These k values are evidence for weak and strong cation interactions in these metal complexes.     

Kinetic studies on partially liganded species of carboxyhemoglobin: (alpha 1CO-beta 1CO)alpha 2 beta 2 or (alpha 2CO beta 2CO)alpha 1 beta 1

Kinetics of CO combination with and dissociation from isomer III, (alpha 1CO beta 1CO)alpha 2 beta 2 or alpha 1 beta 1 (alpha 2CO beta 2CO), and Hb Rothschild have been studied using the double mixing and microperoxidase methods. Isomer III was prepared in a manner so that it was the only reactive species in the reaction mixture. The biphasic reaction time course in both the "on" and "off" reactions of isomer III and the CO combination reaction of Hb Rothschild are attributed to slow relaxation between the fast and slow CO-reacting species in the two proteins: isomer III: l'f = 6 x 10(6) M-1 s-1, l'dimer = 1.7 x 10(6) M-1 s-1, l's = 2.2 x 10(5) M-1 s-1, lf = 0.15 s-1, ls = 0.01 s-1; Hb Rothschild: l'f = 2.8 x 10(6) M-1 s-1; l's = 2.7 x 10(5) M-1 s-1.     

Hemodynamic and proinflammatory actions of endothelin-1 in guinea pig small intestine submucosal microcirculation

The hemodynamic and proinflammatory effects of endothelin-1 (ET-1) in proximal (1st/2nd order) and terminal (3rd/4th order) arterioles and venules were examined in small intestine submucosa of anesthetized guinea pigs. Vessel diameter (D), red blood cell velocity, and blood flow (Q) were determined in eight proximal and eight terminal microvessels before and at 20 min of ET-1 suffusion (10(-10), 10(-9), and 10(-8) M) and then with endothelin-A (ET(A))-receptor blockade with BQ-123 (10(-5) M). This protocol was repeated with platelet-activating factor (PAF) inhibition (WEB-2086, 1.0 mg/kg iv; n = 16). The ET-1-mediated microvascular responses were also examined with endothelin-B (ET(B))-receptor blockade using BQ-788 (10(-5) M; n = 11) alone or with ET(A+B)-receptor blockade with BQ-123 + BQ-788 (n = 10). Microvascular permeability was assessed by FITC-albumin (25 mg/kg iv) extravasation in seven series: 1) buffered modified Krebs solution suffusion (n = 6), 2) histamine suffusion (HIS; 10(-3) M, n = 5), 3) ET-1 suffusion (10(-8) M, n = 5), 4) BQ-123 (10(-5) M) plus ET-1 suffusion (n = 5), 5) PAF inhibition before ET-1 suffusion (n = 5), 6) histamine-1 (H1)-receptor blockade (diphenhydramine, 20 mg/kg iv) before ET-1 suffusion (n = 5), and 7) ET(B)-receptor blockade before (BQ-788 10(-5) M; n = 3) or with ET-1 suffusion (n = 3). D and Q decreased at 10(-8) M ET-1 and returned to control values with BQ-123 and BQ-123+BQ788 but not with BQ-788 in proximal microvessels. D did not change in terminal microvessels with ET-1 (10(-8) M) but decreased with BQ-788 and increased with BQ-123. PAF inhibition did not affect the D and Q responses of proximal microvessels to ET-1 but prevented the fall in Q in terminal microvessels with ET-1. ET-1 increased vascular permeability to approximately 1/3 of that with HIS; this response was prevented with BQ-123 and WEB-2086 but not with H1-receptor blockade. This is the first evidence that submucosal terminal microvessel flow is reduced with ET-1 independent of vessel diameter changes and that this response is associated with increased microvascular permeability mediated via ET(A)-receptor stimulation and PAF activation.     

Significance of ED50 and therapeutic indexes of various radiation-protective agent groups]

Radioprotective agents are divided in 3 groups: (1) cystamine, AET, cystaphos, gammaphos, and thiogammaphos with ED50 (the dose that gives a half of the maximal protective effect) of 10(3)-10(1.6) mumol/kg and therapeutic index K = LD50/ED50 = 10(0)-10(1.6); (2) 5-methoxytryptamine, phenylephrine, serotonin, and norepinephrine with ED50 = 10(1)-10(0) mumol/kg and K = 10(1.8)-10(2,6); (3) clonidine and isoprenaline with ED50 = 10(-0.5)-10(-0.8) mumol/kg and K = 10(3)-10(4). Possible causes of these differences and advantages of low ED50 and high K are discussed.     

Design, synthesis, and biological evaluation of 10-methanesulfonyl-DDACTHF, 10-methanesulfonyl-5-DACTHF, and 10-methylthio-DDACTHF as potent inhibitors of GAR Tfase and the de novo purine biosynthetic pathway

The synthesis and evaluation of 10-methanesulfonyl-DDACTHF (1), 10-methanesulfonyl-5-DACTHF (2), and 10-methylthio-DDACTHF (3) as potential inhibitors of glycinamide ribonucleotide transformylase (GAR Tfase) and aminoimidazole carboxamide ribonucleotide transformylase (AICAR Tfase) are reported. The compounds 10-methanesulfonyl-DDACTHF (1, K(i) = 0.23 microM), 10-methanesulfonyl-5-DACTHF (2, K(i) = 0.58 microM), and 10-methylthio-DDACTHF (3, K(i) = 0.25 microM) were found to be selective and potent inhibitors of recombinant human GAR Tfase. Of these, 3 exhibited exceptionally potent, purine sensitive growth inhibition activity (3, IC50 = 100 nM) against the CCRF-CEM cell line being 3-fold more potent than Lometrexol and 30-fold more potent than the parent, unsubstituted DDACTHF, whereas 1 and 2 exhibited more modest growth inhibition activity (1, IC50 = 1.0 microM and 2, IC50 = 2.0 microM).     

Interaction of pirprofen enantiomers with human serum albumin.

The interaction of pirprofen enantiomers with human serum albumin (HSA) was investigated by means of high-performance liquid chromatography (HPLC), circular dichroism (CD), and 1H NMR spectroscopy. HPLC experiments indicated that both pirprofen enantiomers were bound to one class of high-affinity binding sites (n(+) = 1.91 +/- 0.13, K(+) = (4.09 +/- 0.64) x 10(5) M-1, n(-) = 2.07 +/- 0.13, K(-) = (6.56 +/- 1.35) x 10(5) M-1) together with nonspecific binding (n'K'(+) = (1.51 +/- 0.21) x 10(4) M-1, n'K'(-) = (0.88 +/- 0.13) x 10(-4) M-1). Slight stereoselectivity in specific binding was demonstrated by the difference in product n(+)K(+) = (0.77 +/- 0.08) x 10(6) M-1 vs. n(-)K(-) = (1.30 +/- 0.21) x 10(6) M-1, i.e., the ratio n(-)K(-)/n(+)K(+) = 1.7. CD measurements showed changes in the binding sites located on the aromatic amino acid side chains (a small positive band at 315 nm and a pronounced negative extrinsic Cotton effect in the region 250-280 nm). The protein remains, however, in its predominantly alpha-helical conformation. The 1H NMR difference spectra confirmed that both pirprofen enantiomers interacted with HSA specifically, most probably with site II on the albumin molecule.     

The binding of 6-demethylchlortetracycline to 70S, 50S and 30S ribosomal particles: a quantitative study by fluorescence anisotropy

The binding of demeclocycline (6-demethylchlortetracycline) to ribosomes and ribosomal subunits from Escherichia coli was investigated by using the fluorescence anisotropy of the antibiotic to determine the extent of binding. Binding data obtained from 70S and 30S particles differed fundamentally from those obtained from 50S subunits: the first two showed a strong, specific interaction while the third did not. In addition, all three particles possessed weak, unspecific binding sites. Computer-aided least-squares analysis of the data yielded the following numbers of sites and equilibrium constants: for 30S, n1 = 1, K1 = 2.2 X 10(6) M-1, n2 K2 = 0.029 X 10(6) M-1; for 50S, n1 = 0, n2 K2 = 0.035 X 10(6) M-1; for 70S, n1 = 1, K1 = 3.2 X 10(6) M-1, n2 K2 = 0.082 X 10(6) M-1. These data resolve current disagreement in the literature and are a prerequisite for quantitative studies of the mechanism of inhibition by tetracycline of protein biosynthesis.     

Immunochemical analysis and protective properties of components of Pseudomonas aeruginosa mucus with different molecular weights

P. aeruginosa slime has been separated into fractions XM-300 (3 X 10(5) daltons and more), XM-100 (1 X 10(5) = 3 X 10(5) daltons), PM-30 (3 X 10(4) = 1 X 10(5) daltons) and PM-10, (1 X 10(4) = 3 X 10(4) daltons) by ultrafiltration. The high-molecular slime components (3 X 10(4) daltons and more) have been found to be serologically more active than the low-molecular components (1 X 10(4) = 3 X 10(4) daltons). As shown in experiments on mice, both high-molecular toxic and low-molecular nontoxic slime components have protective activity, but the high-molecular components are more active than the low-molecular ones. The slime components stimulate the formation of immunity to homologous and partially heterologous P. aeruginosa strains in mice. Antigenic relationship between the slime components (especially the high-molecular ones) and the corresponding lipopolysaccharides has been noted.     

The combination of the interleukin-1alpha (IL-1alpha-889) genotype and the interleukin-10 (IL-10 ATA) haplotype is associated with increased interleukin-10 (IL-10) plasma levels in healthy individuals

Family studies have demonstrated striking differences between individuals in their ability to produce IL-10 following lipopolysaccharide (LPS) stimulation of whole blood cultures in vitro, suggesting that differences in IL-10 production involve a considerable hereditary component. The first aim of this study was to analyse the possible effect of IL-10 genotypes and haplotypes on IL-10 plasma levels in a healthy Finnish population. As previous reports have demonstrated that endogenously produced IL-1 induces LPS-stimulated IL-10 production and that IL-10 inhibits synthesis of IL-1 in human monocytes, it is apparent that these two cytokines form an autoregulatory feedback loop. Secondly, we were interested whether any relationship could be found between IL-10 and IL-1beta in vivo. To examine this, the influence of IL-1alpha -889, IL-1beta -511 and IL-1Ra VNTR genotypes and IL-10 genotypes/haplotypes (ACC, GCC and ATA) on IL-10 plasma levels, and a putative correlation between IL-10 and IL-1alpha plasma levels were analysed. Four hundred adult blood samples were obtained from the Finnish Red Cross Blood Transfusion Centre, Tampere. The IL-10, IL-1alpha, IL-1beta and IL-1Ra gene polymorphisms were analysed using PCR. IL-1beta and IL-10 plasma levels were measured using an ELISA method. Our results indicated that increased IL-10 plasma levels were associated with the ATA haplotype (p = 0.03) and, surprisingly, with the IL-1alpha allele 2 carrier status (p = 0.02) in healthy individuals. This IL-1alpha 2+/ATA+ combination was found in 93 subjects out of 400 analysed (23%) and was associated with significantly high IL-10 plasma levels (p = 0.002). When individuals were classified into three groups, with no detectable IL-10 plasma levels (n = 145), with moderate levels (n = 152) and with high levels (n = 100) of IL-10, the IL-1alpha2+/ATA+ combination was more likely present among those with high levels than among those with undetectable levels of IL-10 (OR = 3.3, 95% CI 1.8 - 6.0, p < 0.001) or those with moderate levels of IL-10 (OR = 2.0, 95% CI 1.2 - 3.6, p = 0.012). Besides the observed association between IL-1alpha genotype and IL-10 levels, a moderate correlation was found between IL-10 and IL-1beta levels (r = 0.6, p = 0.01) among IL-10 producers (n = 252). The present findings suggest that the genotype combination of IL-1alpha 2+/ATA+ has a regulatory effect on basal IL-10 levels and that among individuals with measurable IL-10 plasma levels, IL-1beta and IL-10 basal levels correlate. Until now, data on the feedback loop between IL-1 and IL-10 cytokines have been based on studies in vitro, but now our results suggest that this relationship may also exist in vivo.     

In vitro effect of pesticides (dichlofluanid, endosulfan, simazine, tolylfluanid and triallate) on proliferative activity of animal derived cell cultures

In this study pesticides with different chemical structures (dichlofluanid, endosulfan, simazine, tolylfluanid and triallate) were examined for their potential cytotoxic effect on proliferative activity of cell cultures of mammalian origin. Cell lines Madin-Darby Bovine Kidney (MDBK), Rabbit Kidney (RK13), Porcine Kidney (PK15), and semicontinual line of Bovine Embryonic Pulmonary Cells (BEPC) were used in the study. From these cell cultures cell proliferative activity was suppressed most intensively in PK15 culture by endosulfan (10(-1) - 10(-6) M). The least effect on cell proliferation in all cell cultures tested, with the exception PK 15 (10(-1) - 10(-2) M), was recorded after simazine exposure. On the basis of IC50 values the cytotoxic effect was: dichlofluanid (IC50 = 10(-3.94) M) > tolylfluanid (IC50 = 10(-3.69) M) > endosulfan (IC50 = 10(-3.24) M) > triallate (IC50 = 10(-3.12) M) > simazine (IC50 = 10(-1.78) M). The comparison of average IC50 values of cell cultures revealed that the most sensitive cell lines were PK15 (IC50 = 10(-3.27) M) and RK13 (IC50 = 10(-3.21) M), whereas MDBK (IC50 = 10(-2.55) M) and BEPC (IC50 = 10(-2.52) M) were less sensitive to pesticide exposure.     

Binding of branched-chain 2-oxo acids to bovine serum albumin.

1. Binding of branched-chain 2-oxo acids to defatted bovine serum albumin was shown by gel chromatography and equilibrium dialysis. 2. Equilibrium-dialysis data suggest a two-side model for binding in Krebs-Henseleit saline at 37 degrees C with n1 = 1 and n2 = 5. Site association constants were: 4-methyl-2-oxovalerate, k1 = 8.7 x 10(3) M-1, k2 = 0.09 x 10(3) M-1; 3-methyl-2-oxovalerate, k1 = 9.8 x 10(3) M-1, k2 = 0.08 x 10(3) M-1; 3-methyl-2-oxobutyrate, k1 = 1.27 x 10(3) M-1, k2 = less than 0.05 x 10(3) M-1. 3. Binding of 4-methyl-2-oxovalerate to defatted albumin in a phosphate-buffered saline, pH 7.4, gave the following thermodynamic parameters: primary site delta H0(1) = -28.6kJ . mol-1 and delta S0(1) = -15.2J . mol-1 . K-1 (delta G0(1) = -24.0kJ . mol-1 at 37 degrees C) and secondary sites delta H0(2) = -25.4kJ . mol-1 and delta S0(2) = -46.1J . mol-1 . K-1 (delta G0(1) = -11.2kJ . mol-1 at 37 degrees C). Thus binding at both sites is temperature-dependent and increases with decreasing temperature. 4. Inhibition studies suggest that 4-methyl-2-oxovalerate may associate with defatted albumin at a binding site for medium-chain fatty acids. 5. Binding of the 2-oxo acids in bovine, rat and human plasma follows a similar pattern to binding to defatted albumin. The proportion bound in bovine and human plasma is much higher than in rat plasma. 6. Binding to plasma protein, and not active transport, explains the high concentration of branched-chain 2-oxo acids leaving rat skeletal muscle relative to the concentration within the tissue, but does not explain the 2-oxo acid concentration gradient between plasma and liver.     

Reversible, slow, tight-binding inhibition of human leukocyte elastase

CBz-Ala-Ala-Pro-ambo-Val-CF3 (1) was synthesized. The compound inhibits human Leucocyte elastase with Ki = 1.0 x 10(-9) M. This inhibitor is reversible, slow, tight-binding inhibitor with k on = 2 x 10(4) M-1 s-1 k off = 1.9 x 10(-5) s-1. For the solubilization of elastin by HLE by 1 I.C. 50 = 110 nM. This inhibitor is the most effective aldehyde or ketone inhibitor of a serine proteinase yet described.