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Measurement of the bioactivity of interleukin and tumour necrosis factor in synovial fluid of Kashin-Beck disease.

We studied the bioactivity of interleukin (IL-1) and tumour necrosis factor (TNF) in the synovial fluid of 12 patients with Kashin-Beck disease (KBD), 8 patients with osteoarthritis (OA) and 8 normal controls. C57 female mouse T cell proliferation method was used to test the bioactivity of IL-1, and lethality method with L929 cells to test the bioactivity of TNF. The bioactivities of IL-1 and TNF in the synovial fluid from patients with KBD were higher than those in the synovial fluid of osteoarthritis and normal controls, indicating the participation of IL-1 and TNF in the pathogenesis of KBD.

Adult↗

Sustainable production of bioactive compounds by sponges--cell culture and gene cluster approach: a review.

Sponges (phylum Porifera) are sessile marine filter feeders that have developed efficient defense mechanisms against foreign attackers such as viruses, bacteria, or eukaryotic organisms. Protected by a highly complex immune system, as well as by the capacity to produce efficient antiviral compounds (e.g., nucleoside analogues), antimicrobial compounds (e.g., polyketides), and cytostatic compounds (e.g., avarol), they have not become extinct during the last 600 million years. It can be assumed that during this long period of time, bacteria and microorganisms coevolved with sponges, and thus acquired a complex common metabolism. It is suggested that (at least) some of the bioactive secondary metabolites isolated from sponges are produced by functional enzyme clusters, which originated from the sponges and their associated microorganisms. As a consequence, both the host cells and the microorganisms lost the ability to grow independently from each other. Therefore, it was--until recently--impossible to culture sponge cells in vitro. Also the predominant number of "symbiotic bacteria" proved to be nonculturable. In order to exploit the bioactive potential of both the sponge and the "symbionts," a 3D-aggregate primmorph culture system was established; also it was proved that one bioactive compound, avarol/avarone, is produced by the sponge Dysidea avara. Another promising way to utilize the bioactive potential of the microorganisms is the cloning and heterologous expression of enzymes involved in secondary metabolism, such as the polyketide synthases.

Amino Acid Sequence↗

Arachidonic acid-derived bioactive lipids: their role and the role for their inhibitors in dermatology.

BACKGROUND: In addition to corticosteroids, there are increasing numbers of anti-inflammatory agents that specifically target bioactive lipids generated from arachidonic acid. Knowledge of the diverse mechanisms of action of these different bioactive lipids holds promise in the therapy of a wide spectrum of cutaneous and systemic disorders. OBJECTIVE: Therapeutic manipulations of these lipid molecules through inhibition, stimulation, or direct replacement have broad physiologic effects. These therapeutic strategies not only modulate inflammation, pain, and hemostatic parameters, they also play a role in cardiac, respiratory, renal, and gastrointestinal function and disease, as well as in angiogenesis and in factors that control cell growth and apoptosis important in carcinogenesis. CONCLUSION: Newer drug discovery methods, including combinatorial chemistry with molecular modeling, have made it possible to develop inhibitors and analogs with increasing specificity and bioactivity and decreasing toxicity. Although the application of these analogs and inhibitors for cutaneous disease is limited today, either as primary agents or adjuvant therapy, these drugs will have a place in our therapeutic regimes of the future. We present a review of the therapeutic agents now available from manipulation of these bioactive lipids, and their role and future in dermatology.

Anti-Inflammatory Agents↗

Microstructural and in vitro characterization of SiO2-Na2O-CaO-MgO glass-ceramic bioactive scaffolds for bone substitutes.

In the present research work, the preparation and characterization of bioactive glass-ceramic scaffolds for bone substitutes are described. The scaffolds were prepared by starch consolidation of bioactive glass powders belonging to the SiO2-Na2O-CaO-MgO system using three different organic starches (corn, potatoes and rice) as reported in a previous screening process. The scaffolds, characterized by scanning electron microscopy, showed a porous structure with highly interconnected pores. The pores sizes assessed by mercury intrusion porosimetry put in evidence the presence of pores of 50-100 microm. The structure of the scaffolds was investigated by X-ray diffraction and revealed the glass-ceramic nature of the obtained material. The mechanical properties of the scaffolds were evaluated by means of compressive tests on cubic samples and the obtained results demonstrated their good mechanical strength. The in vitro bioactivity of the scaffolds was tested by soaking them in a simulated body fluid (SBF) and by subsequently characterizing the soaked surfaces by SEM, EDS and X-ray diffraction. Good in vitro bioactivity was found for the starting glass and for the obtained scaffolds. Moreover, the scaffold bioresorption, tested by measuring the samples weight loss in SBF at different periods of time, showed a partial resorption of the scaffolds. Cell culture testing of the three different scaffolds indicated no differences in cell number and in alkaline phosphatase activity; the morphology of the osteoblasts showed good spreading, comparable to bulk material which was used as the control.

Body Fluids↗

Gene activation by bioactive glasses.

Bioactive glasses have been shown to regulate gene expression in both hard and soft tissue repair. New resorbable bioactive glass constructs are now being developed that can influence gene expression in the local environment by manipulating material properties such as the surface chemistry, topography and the release of dissolution ions. The success of these scaffolds, however, may depend upon a greater understanding of the bioactive glass stimulated gene expression pathways. This will allow the construction of tissue specific scaffolds with tailored surface chemistry, topography and ion release rates. This paper summarises the advances made in understanding gene expression in response to bioactive glasses and discusses the future steps required for further insights into these molecular mechanisms.

Animals↗

From the bioactive glasses to the star gels.

The improvement of bioactive glasses is one of the most important subjects in the field of hard tissue replacement. More than 30 years after being discovered by Prof. Hench, bioactive glasses still attract the attention of many researchers all over the World. With this article we want to pay homage to Prof. Hench by means of reviewing the main contribution of our research team to the field initiated by him in 1969. Our efforts, aimed to go further in the understanding of sol-gel glasses bioactivity as well as to improve the mechanical properties of bioactive materials, are explained.

Biocompatible Materials↗

Swelling properties and bioactivity of silica gel/pHEMA nanocomposites.

A novel hydrogel based on 2-hydroxyethyl- methacrilate and SiO(2) nanoparticles was prepared. The filler was added at a concentration of 30% w/w of silica nanoparticles to the mass of polymer. The composite material was characterised as far as concerns swelling behaviour in comparison to pHEMA. Swelling ratio of modified pHEMA was higher. Bioactivity of both SiO(2) nanoparticles and the modified hydrogel was evaluated by soaking samples into a simulated body fluid (SBF). FT-IR spectroscopy, scanning electron microscopy (SEM) and energy dispersive system (EDS) results suggest silica nanoparticles keep bioactive in the polymer. SiO(2) filler in a p(HEMA) matrix makes the composite bioactive. Therefore, these composites can be used to make bioactive scaffold for bone engineering.

Absorption↗

Behaviour of MG-63 osteoblast-like cells on wood-based biomorphic SiC ceramics coated with bioactive glass.

The aim of this study was to test the in vitro cytotoxicity of wood-based biomorphic Silicon Carbide (SiC) ceramics coated with bioactive glass, using MG-63 human osteoblast-like cells, with a view to their application in bone implantology. To better understand the scope of this study, it should be taken into account that biomorphic SiC ceramics have only recently been developed and this innovative product has important properties such as interconnected porosity, high strength and toughness, and easy shaping. In the solvent extraction test, all the extracts had almost no effect on cellular activity even at 100% concentration, and cells incubated in the bioactive glass-coated SiC ceramics extracts showed a proliferation rate similar to that of the Thermanox control. There were no significant differences when the cellular attachment response of the cells on the wood-based biomorphic SiC ceramics, uncoated or coated with bioactive glass, was compared to the one exhibited by reference materials like Ti6Al4V and bulk bioactive glass. This fact looks very promising for biomedical applications.

Biomimetic Materials↗

Spontaneously resorbing ectopic pregnancy: preservation of human chorionic gonadotropin bioactivity despite declining steroid hormone levels.

We compared serum hormone profiles of patients with normal intrauterine pregnancies (n = 14), spontaneously resorbing ectopic pregnancies (n = 10), and viable ectopic pregnancies (n = 26). Hormone profiles were evaluated at 5 to 8 weeks' gestational age. Ectopic pregnancies were diagnosed by laparoscopy; intrauterine gestations were confirmed by ultrasonography. Immunoreactive beta-human chorionic gonadotropin, progesterone, estradiol, and 17-hydroxyprogesterone were measured by radioimmunoassay. Bioactive human chorionic gonadotropin was measured by a modified mouse Leydig cell bioassay. Diminished steroid production was noted in ectopic pregnancies; levels in serum of patients with resorbing ectopic pregnancies were lower than values expressed in viable ectopic pregnancies (p less than 0.01). Serum levels of human chorionic gonadotropin bioactivity correlated closely with immunoreactive human chorionic gonadotropin in all three groups (r = 0.81, p less than 0.01). Ratios of bioactive human chorionic gonadotropin to immunoreactive beta-human chorionic gonadotropin were similar (0.93 +/- 0.26 in resorbing ectopic pregnancies, 1.11 +/- 0.16 in viable ectopic pregnancies, and 0.90 +/- 0.10 in intrauterine pregnancies). We conclude that although reduced serum levels of steroids noted in ectopic pregnancy suggest an impairment in corpus luteum activity, diminished steroid production was not attributable to lower human chorionic gonadotropin bioactivity.

17-alpha-Hydroxyprogesterone↗

Synthesis and in vitro bioactivity of C-terminal deleted analogs of human growth hormone-releasing factor.

A series of C-terminal deleted analogs of human growth hormone-releasing factor (hGRF) with either an amidated or a free carboxylic acid C-terminus were synthesized by solid phase methodology. Their capacity to release growth hormone was tested on rat anterior pituitary cells in monolayer culture. A gradual decrease of bioactivity down to 23% relative to hGRF was noted when the C-terminal amino acids were deleted to hGRF (1-34)OH. Further deletions, however, did not decrease the bioactivity because the potencies of the fragments, hGRF(1-31)NH2, (1-30)NH2 and (1-29)NH2 remained at about 50% of that of hGRF. Continual deletion of residues to hGRF(1-23)NH2, (1-22)NH2 and (1-21)NH2 still yielded bioactive fragments with full intrinsic activity despite very low potency. Only with the deletion down to hGRF(1-19)NH2 did the bioactivity completely disappear. Thus, together with the data published in a previous paper (1), the minimal biologically active core of hGRF with full intrinsic activity comprises the fragment (3-21).

Animals↗

Bioactivation of 4-fluorinated anilines to benzoquinoneimines as primary reaction products.

Metabolism and bioactivation of fluoroanilines was studied both in vitro in microsomal systems and in vivo. 4-Fluoroaniline and pentafluoroaniline and their non-para fluorinated analogues were used as the model compounds. Special attention was focussed on bioactivation to reactive benzoquinoneimines. Cytochrome P-450 dependent monooxygenation at a non-fluorinated para position in (fluoro)aniline derivatives proceeds by formation of the para hydroxylated derivative as the primary metabolite. Monooxygenation at a fluorinated para position in an aniline derivative, however, proceeds by formation of fluoride anion and the reactive benzoquinoneimine as primary reaction products. Thus, for fluoroanilines with a fluorine substituent at the para position bioactivation to the reactive benzoquinoneimine can be a direct result of the cytochrome P-450 dependent conversion. In systems containing NAD(P)H and/or other reducing equivalents part of the benzoquinoneimine can be chemically reduced to give the corresponding 4-hydroxyaniline. In vivo this reduced form of the metabolite can be sulphated or glucuronidated and excreted into urine. The results obtained point to increased chances of bioactivation for aniline derivatives with a fluorinated para position as compared to their non-para fluorinated analogues, both in vitro but also in vivo.

Aniline Compounds↗

Extrahepatic bioactivation of aflatoxin B1 in fetal, infant and adult rats.

Whole-body autoradiography of 3H-labelled aflatoxin B1 (3H-AFB1) in female non-pregnant adult and infant Sprague-Dawley rats showed retention of tissue-bound radioactivity, in addition to the liver, in the mucosa and some glands in the nose, and in the mucosa of the nasopharynx, trachea, bronchioles, colon and caecum. The extrahepatic binding was most pronounced in the infant rats. In a rat pretreated with the glutathione (GSH)-depleting agent phorone, bound labelling was also seen in the superficial part of the mucosa of the glandular stomach. Autoradiography of 3H-AFB1 in pregnant rats showed a marked localization of bound AFB1-metabolites in the fetal nasal olfactory and tracheal mucosa. In vitro experiments demonstrated that the nasal olfactory mucosa had a much higher capacity than the liver to form AFB1-metabolites which bound to DNA and protein. The bioactivation was observed both pre- and post-natally and increased with age. Bioactivation was found also in the caecum, the colon and the lateral nasal gland (Steno's gland), but not in the small intestine, oesophagus or Harderian gland. Our results indicated that glutathione-S transferase activity catalysing the AFB1-8,9-epoxide GSH-conjugation was present in the nasal olfactory mucosa and liver at all pre- and post-natal ages examined. Several of the extrahepatic tissues able to bioactivate AFB1 have been reported to be targets for the carcinogenicity of the substance. Our results indicate that the extrahepatic carcinogenicity of AFB1 is correlated to a local bioactivation in the sensitive tissues.

Aflatoxin B1↗

Bioactivation of catechol in rat and human bone marrow cells.

o-Benzoquinone-glutathione (GSH) conjugate formation and covalent binding of [14C]catechol to protein were utilized as probes of bioactivation of catechol in both rat and human white bone marrow cell systems. Conjugate formation and binding occurred in the absence of exogenous hydrogen peroxide, but were markedly stimulated by its addition. Protein-binding and conjugate formation using rat cells in the presence of exogenous peroxide were increased by the presence of phenol whereas GSH and hydroquinone inhibited binding. Similarly, protein-binding in the absence of exogenous peroxide was inhibited by GSH and exacerbated by phenol. Prostaglandin synthase, the peroxidatic function of which may also utilize hydrogen peroxide as a substrate, appeared on the basis of experiments using arachidonic acid to play only a minor role in bioactivation of catechol in rat bone marrow cells. These results show that peroxide-dependent bioactivation of catechol occurs in rat and human bone marrow cells and that hydroquinone and GSH inhibit whereas phenol stimulates bioactivation.

Animals↗

Metabolic evidence for the involvement of enzymatic bioactivation in the cataractogenicity of acetaminophen in genetically susceptible (C57BL/6) and resistant (DBA/2) murine strains.

Acetaminophen has been shown to be cataractogenic in mice and rabbits. C57BL/6 and DBA/2 mice respectively are genetically susceptible and resistant to the induction of cytochrome P-448 by 3-methylcholanthrene (3-MC). This isoenzyme is thought to bioactivate acetaminophen to a toxic reactive intermediate. These two murine strains also are correspondingly susceptible and resistant to acetaminophen cataractogenesis. To evaluate the potential role of enzymatic bioactivation as a determinant of acetaminophen cataractogenesis, C57BL/6 and DBA/2 mice were treated with acetaminophen, 300 or 400 mg/kg intraperitoneally (ip), with or without pretreatment 48 hr earlier using 3-MC, 200 mg/kg ip. Lenticular cataracts were evaluated using the unaided eye and a slit lamp, and hepatotoxicity was evaluated by determination of peak plasma concentration of alanine aminotransferase (ALT). Plasma concentrations of acetaminophen and metabolites, particularly the glutathione (GSH)-derived conjugates (cysteine and mercapturic acid) reflecting enzymatic bioactivation, were measured by high-performance liquid chromatography. Cataracts developed only in C57BL/6 mice pretreated with 3-MC, occurring in 1 of 5 and 5 of 5 animals treated respectively with 300 and 400 mg/kg of acetaminophen. Comparing these two groups of induced C57BL/6 mice, production of the cysteine conjugate of acetaminophen was 2.5-fold higher with the 400 mg/kg dose of acetaminophen (p less than 0.05). Compared to their respective dose-matched, noninduced controls, cysteine conjugate production in the 300 and 400 mg/kg dose groups of induced C57BL/6 mice respectively was 3-fold and 4-fold higher (p less than 0.05). No DBA/2 mice developed cataracts. No mercapturic acid conjugate was detectable in the plasma of DBA/2 mice, and production of the cysteine conjugate was not altered in this strain by increasing the dose of acetaminophen or by pretreatment with 3-MC. The mean peak plasma concentration of the cysteine conjugate, reflecting acetaminophen bioactivation, was 5-fold higher in animals developing cataracts compared with those without cataracts (p less than 0.001). Plasma concentrations of unmetabolized acetaminophen were similar in all groups and unrelated to the development of cataracts. All mice of both strains pretreated with 3-MC showed evidence of hepatotoxicity, indicating a dissociation between hepatotoxic and cataractogenic susceptibility.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetaminophen↗

In vitro observations of iron-doped bioactive glasses.

Several in vitro tests are necessary to evaluate the biological compatibility of materials used to manufacture implants for humans or animals. These tests will indicate whether these materials can be tolerated when implanted in vivo. Since ceramic materials are suitable for coating layers, it was decided to study their performance by adding certain oxides as doping agents. This paper describes their influence in vivo in some preliminary tests and in vitro for iron trioxide only. This work attempts to ascertain the range of doping to maintain the bioactivity characteristics and in particular the influence of iron trioxide as a doping agent in the bioactive glass. To do so the adhesion of baby hamster kidney cells to bioactive glasses doped with different concentrations (2 and 4 wt%) of this compound was quantified. The base glass without iron oxide was used as a control as 0 wt% doping. In a second phase, a coating of fibronectin, a glycoprotein that plays an important role in cell binding and spreading on substrates, was quantified on the same surfaces. It was shown that the cell avidity for the surfaces of the materials increases with increased doping of iron oxide. The values are significantly higher than those observed for the base glass. Fibronectin adsorption quantification showed a similar trend, with values of adsorption similar for the doped samples and higher than that of the non-doped glass. It was demonstrated that iron trioxide induces an increase in cell adhesion capability with bioactive glass surfaces and the capability of adhesion of the fibronectin to act as a substrate on the same surfaces.

Adsorption↗

Improved high bioactivation cross for the wing somatic mutation and recombination test in Drosophila melanogaster.

The two tester strains of the high bioactivation (HB) cross for the wing somatic mutation and recombination test (SMART) in Drosophila melanogaster developed by Frölich and Würgler possess high metabolic capacity to activate promutagens. These strains contain chromosomes 1 and 2 of the DDT-resistant stock Oregon R(R) which exhibits a high constitutive level of cytochrome P450. However, they show several disadvantages for routine application, such as disturbed wing hair patterns in certain areas of the wing, making spot classification difficult, and a delay in development of the larvae. We have established and evaluated an improved HB cross (ORR; flr3 females and mwh males) producing ORR heterozygous individuals. These develop normally and have a normal, undisturbed wing hair pattern while exhibiting high bioactivation. The hybrid larvae of the improved HB cross show P450-dependent bioactivation capacity equal to or even slightly higher than those of the original HB cross. This was demonstrated by measuring the genotoxic activity of the promutagens diethylnitrosamine, 7,12-dimethylbenz[a]anthracene, N-nitrosopyrrolidine, and urethane. In addition, the improved HB cross has a sensitivity to the direct-acting alkylating agent ethyl nitrosourea equal to that of the standard cross. The main advantage of the improved HB cross is to combine the high bioactivation capacity with the ease of scoring the wings using the same criteria as for the standard cross.

Animals↗

Follicle-stimulating hormone-stimulated secretion of an immunoreactive 29 kDa inhibin alpha-subunit complex, but not of 32 kDa bioactive inhibin, from cultured immature rat Sertoli cells.

The medium of cultured Sertoli cells from immature rat testes contains 29 and 32 kDa proteins, which are recognized by an antiserum against the 22 N-terminal amino acids of the inhibin alpha-subunit. These proteins were detected by immunoprecipitation of labelled proteins after incubation of Sertoli cells with [35S]methionine, and by Western blotting. The amount of the 32 kDa protein was not affected by the addition of follicle-stimulating hormone (FSH) to the culture medium of the Sertoli cells, whereas FSH induced a large increase of the amount of the 29 kDa protein. Finally, the 29 and 32 kDa proteins in the medium from control and FSH-stimulated Sertoli cells were separated by sodium dodecyl sulfate-polyacrylamide electrophoresis, and inhibin bio- and immunoactivity were determined in eluates of the slices of the gel. Equal amounts of bioactivity were found in control and FSH-stimulated samples at 32 kDa, while the amount of immunoactivity at 29 kDa was increased; no bioactivity was detected in the eluates of these slices. It is concluded that FSH stimulates the secretion of a 29 kDa inhibin-like protein, which does not contain inhibin bioactivity. This indicates that results of experiments, in which antibodies against N-terminal peptides of the inhibin alpha-subunit are used to detect inhibin, do not necessarily reflect the amount of bioactive inhibin produced.

Animals↗

Mitochondrial bioactivation of cysteine S-conjugates and 4-thiaalkanoates: implications for mitochondrial dysfunction and mitochondrial diseases.

The toxicity of most drugs and chemicals is associated with their enzymatic conversion to toxic metabolites. Bioactivation reactions occur in a range of organs and organelles, including mitochondria. The toxicity of haloalkene-derived cysteine S-conjugates and related 4-thiaalkanoates is associated with their mitochondrial bioactivation. Toxic cysteine S-conjugates are formed by the glutathione S-transferase-catalyzed addition of glutathione to haloalkenes to give glutathione S-conjugates, which are hydrolyzed by gamma-glutamyltransferase and dipeptidases. Mitochondrial cysteine conjugate beta-lyase-catalyzed bioactivation of cysteine S-conjugates affords unstable alpha-halothiolates. Haloalkene-derived 4-thiaalkanoates, which are analogs of cysteine S-conjugates that lack an alpha-amino group, undergo bioactivation by the enzymes of fatty acid beta-oxidation to give 3-hydroxy-4-thiaalkanoates that eliminate alpha-halothiolates. alpha-Halothiolates yield alkylating and acylating agents that interact with cellular macromolecules and thereby cause cell damage. Mitochondrial dysfunction is the hallmark of cysteine S-conjugate-induced cytotoxicity: decreased respiration, decreased ATP and total adenine nucleotide concentrations, depletion of the mitochondrial glutathione content, perturbations in cellular Ca2+ homeostasis, and damage to the mitochondrial genome are seen with cysteine S-conjugates. Similar changes are observed with cytotoxic 4-thiaalkanoates, but inhibition of the medium-chain acyl-CoA dehydrogenase and hypoglycemia are also observed.

Animals↗