Ultrastructural effects of silicic acid on primary lung fibroblasts in tissue culture

Transmission and scanning electron microscopic examination of primary lung fibroblasts exposed in tissue culture to polymeric silicic acid (PSA) revealed profound cellular changes in the cell surface membranes, resulting in rapid endocytosis of affected membranes and formation of multivesicular bodies. Exposure to monomeric silicic acid did not appear to exhibit any immediate adverse effects. Appearance of numerous cytoplasmic vacuoles within 1 h of PSA exposure was easily visible by light microscopy. Electron microscopy revealed that PSA exposure caused formation of an 'osmiophilic' cell surface membrane. Numerous osmiophilic cytoplasmic blebs on the surface and subsequent endocytotic vesicles appeared to collapse and aggregate into multivesicular bodies. This study provides ultrastructural evidence of the direct interaction between lung fibroblasts and polymeric silicic acid, which has a dramatic effect the surface membrane, its subsequent internalization and cytoplasmic processing. This interaction could be one of the key steps in the damaging effects of silica containing dust.     

Diatom mineralization of silicic acid

Silicic acid binding, possibly at a membrane site is suggested as the first step in Si(OH)₄ metabolism in the marine diatom Nitzschia angularis. We estimate 1738±133 Si(OH)₄ binding sites·m^-2 cell surface and a turnover of 4–12 Si(OH)₄ molecules·s^-1 at maximum transport velocity. All studies were carried out using ⁶⁸Ge(OH)₄ as a tracer for Si(OH)₄.After Si(OH)₄ uptake, rapid (<1 min) transformation or intracellular binding was demonstrated; yet the extractable pool size after 1 min was indicative of transport against a concentration gradient. Initial uptake kinetics were linear for 150 s and saturation kinetics were demonstrated with kinetic parameters of 560 pmol Si(OH)₄·10⁶ cells^-1·min^-1 (V _max) 4.2 mol Si(OH)₄·L^-1(K_s). A metabolic energy requirement for transport was suggested by inhibition of uptake by agents that uncouple or inhibit phosphorylation; transport also was sensitive to agents that block sulfhydryl groups. Such characteristics are consistent with Si(OH)₄ transport being an active carrier mediated process.Rates of Si(OH)₄ transport were regulated during various growth stages and during the synchronized cell cycle. In Si(OH)₄ starved cells, blocked at the initiation of silica frustule formation, cycloheximide treatment caused a rapid decline of transport rate. Blocked cells, placed in the dark, maintained high transport rates for 6 h after which there was a loss of activity during the following 18 h. Light dependent recovery of transport ability in 12 h predarkened cells was dependent on de novo protein synthesis.Abbreviations ASW artificial sea water - CCCP carbonyl cyanide-m-chlorophenyl hydrazone - DCMU 3(3,4-dichlorophenyl-1,1-dimethyl urea) - 2,4-DNP 2,4-dinitrophenol - ESWT enriched sea water with tryptone - NEM n-ethylmaleimide - pCMB p-chloromercuribenzoic acid     

Cell-cycle effects on the kinetics of silicic acid uptake and resource competition among diatoms

Current models of nutrient utilization by planktonic algae generallyassume that cells transport nutrients continuously. This assumptionis violated in the case of Si utilization by diatoms. Silicicacid transport is confined to specific portions of the cellcycle such that only a fraction of the cells in a populationare likely to be transporting silicic acid at any given time.A theoretical framework for describing the growth and nutrient-uptakekinetics of populations of cells exhibiting cell-cycle-dependentnutrient uptake is presented. Thalassiosira weissflogii wasused as a model organism. Kinetic parameters determined at steadystate, assuming cells take up Si continuously, underestimatedboth the V_m and K_s of individual cells by nearly an order ofmagnitude. Likewise, kinetic constants determined from short-termexperiments assuming continuing uptake were shown to be dependenton the initial physiological state of the population and todramatically underestimate the kinetic parameters of individualcells. The response of steady-state populations to large pulsesof dissolved Si was also examined. The ratio of the averagemaximum specific uptake rate of the culture to the average maximumobservable specific growth rate (V' _mavg/     

Inhibitory effects of sulfamic acid on three thiosulfate-oxidizing chemolithotrophs

Sulfamate is an analogue of thiosulfate, and the sodium and potassium salts of sulfamic acid inhibited the chemolithoautotrophic growth on thiosulfate of Acidithiobacillus ferrooxidans and Halothiobacillus neapolitanus. The chemo-organotrophic growth of Paracoccus versutus on sucrose was similarly inhibited by sulfamate. Thiosulfate oxidation by suspensions of H. neapolitanus was, however, unaffected by sulfamate, showing that sulfamate did not directly affect thiosulfate uptake, activation or oxidation. Inhibition of P. versutus was not relieved by cysteine and methionine, indicating that sulfate uptake and sulfur amino acid biosynthesis were not directly affected by sulfamate. Sulfamate was not degraded by any of the bacteria, and so could not serve as an alternative to thiosulfate as an energy-yielding substrate. Sulfamate is also an analogue of ammonia and might act like hydrazine by inhibiting ammonium uptake or an essential enzyme activity.     

Inhibitory effects of acetylsalicylic acid on exocrine pancreatic carcinogenesis

Abstract

We investigated short (6 months) and long (12 months) term inhibitory effects of low (200 ppm) and high (400 ppm) dosages of acetylsalicylic acid (aspirin) on exocrine pancreatic carcinogenesis. It is known that exocrine pancreatic carcinogenesis can be detected by the presence of atypical acinar cell foci (AACF) in pancreas. We investigated possible inhibitory effects of acetylsalicylic acid in an azaserine-treated rat model. AACF were produced in rats by injection with azaserine according to previous studies. Our findings showed that the number, volume and diameter of pancreatic AACF were reduced after acetylsalicylic acid application. These observations suggest that acetylsalicylic acid may exert a protective effect against neoplastic development of pancreatic acinar cells in azaserine injected rats. Our findings corroborate reports in the literature concerning the effects of aspirin in reducing neoplastic development.     

Inhibitory effects of flavonoids on animal fatty acid synthase

The inhibitory effects of 15 flavonoids on animal fatty acid synthase (FAS, EC 2.3.1.85) were investigated, and 9 of them were found to inhibit FAS with IC(50) (the inhibitor concentration inhibiting 50% of the activity of FAS) values ranging from 2 to 112 microM. A structure-activity relationship study showed that the flavonoids containing two hydroxyl groups in the B ring and 5,7-hydroxyl groups in the A ring in combination with a C-2, 3 double bond were the most inhibitory. Morin (IC(50) = 2.33 +/- 0.09 microM) was further investigated kinetically to detail the inhibitory mechanism. The results showed that morin inhibited the overall reaction of FAS competitively with Ac-CoA, noncompetitively with Mal-CoA and in a mixed manner with NADPH. The study indicated that morin bound reversibly to the beta-ketoacyl synthase domain of FAS to inhibit the elongation of the saturated acyl groups in fatty acids synthesis.     

Degradation of DNA by silicic acid

S1 nuclease hydrolysis and hydroxyapatite chromatography were used to study the effect of silicic acid on DNA. Native calf thymus DNA was incubated with increasing concentrations of silicic acid (DNA nucleotide/silicic acid molar ratios of 1:0.25, 1:0.5 and 1:1) and subjected to S1 nuclease hydrolysis. An increasing degree of DNA degradation was seen suggesting a destabilization of the secondary structure. A decrease in melting temperature was also observed. Hydroxyapatite chromatography indicated that incubation at the molar ratio of 1:1 resulted in denaturation and degradation of DNA.     

Inhibitory effects of gallic acid and quercetin on UDP-glucose dehydrogenase activity

We have examined polyphenols as potential inhibitors of UDP-glucose dehydrogenase (UGDH) activity. Gallic acid and quercetin decreased specific activities of UGDH and inhibited the proliferation of MCF-7 human breast cancer cells. Western blot analysis showed that gallic acid and quercetin did not affect UGDH protein expression, suggesting that UGDH activity is inhibited by polyphenols at the post-translational level. Kinetics studies using human UGDH revealed that gallic acid was a non-competitive inhibitor with respect to UDP-glucose and NAD⁺. In contrast, quercetin showed a competitive inhibition and a mixed-type inhibition with respect to UDP-glucose and NAD⁺, respectively. These results indicate that gallic acid and quercetin are effective inhibitors of UGDH that exert strong antiproliferative activity in breast cancer cells.     

Kinetic study of acid depolymerization of chitosan and effects of low molecular weight chitosan on erythrocyte rouleaux formation

In this study, the depolymerization of chitosan was carried out in an acetic acid aqueous solution and was followed by viscometry for molecular weight determination. It was found that the depolymerization rate increased with elevated temperatures and with high acid concentrations. Based on FTIR analysis, the chitosan was depolymerized randomly along the backbone; no other structural change was observed during the acid depolymerization process. Revealed in the TGA study, the degradation temperature and char yield of LMWCs (low molecular weight chitosan) were molecular weight dependent. The blood compatibility of LMWCs was also investigated: rouleaux formation was observed when erythrocyte contacted with LMWCs, which showed that LMWCs are able to interfere with the negatively charged cell membrane through its polycationic properties. Furthermore, as regards a kinetics investigation, the values of M_n (number-average molecular weight) were obtained from an experimentally determined relationship. The kinetics study showed that the complex salt, formed by amine on chitosan and acetic acid, acted as catalyst. Finally, the activation energy for the hydrolysis of the glycosidic linkage on chitosan was calculated to be 40 kJ/mol; the mechanism of acid depolymerization is proposed. In summary, LMWCs could be easily and numerously generated with acid depolymerization for further biological applications.     

Inhibitory effect of water-soluble chitosan on growth of Streptococcus mutans

The present study was undertaken to evaluate the effects of pH and the degree of polymerization of chitosan on the inhibition of growth of Streptococcus mutans. Three types of chitosan, polymer, oligomer and monomer, were used at 4% (W/V) and three different levels of pH: 6.0, 6.5 and 7.4. Bactericidal activity was calculated by the growth ratio. Chitosan oligomer significantly inhibited bacterial growth at a pH value of 6.5. All three types of chitosan strongly inhibited bacterial growth at pH 6.0. Furthermore, nearly complete inhibition was obtained with 2%(W/V) chitosan solution at constant pH 6.5. This study is the first to report that water-soluble chitosan directly suppresses the growth of the typical cariogenic bacterium S. mutans even at pH 6.5, without causing demineralization of the tooth surface.