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The formation of intestinal villi (organogenesis phase) may be studied in organ culture with a completely synthetic medium in 15-day fetal mouse duodenal explants. However, in these explants absorptive cells remained poorly differentiated with all the hormones studied except with epidermal growth factor. In order to elucidate the role of hormones and other factors on the maturation of absorptive cells (maturation phase) in the fetal rodent in organ culture, we have taken the explants after the organogenesis phase. We have studied different culture conditions and have found that 17-day mouse duodenal explants can be cultured during 48 hours with Leibovitz L-15 medium in a 95% O2-5% CO2 atmosphere provided that the explants are relatively large (5 X 2 mm). With this method, dexamethasone (Dx) has been shown to have a direct effect on the maturation of the fetal duodenal mucosa. The addition of Dx (300 ng/ml) to the completely synthetic medium 1) improves the morphology of the explants, 2) induces a significant increase in maltase activity in the tissues, and 3) reduces significantly the labeling index of the duodenal explants after 48 hours of culture. Direct action of Dx on the duodenal mucosa is shown for the first time in organ culture using a completely synthetic medium. This method will permit us to study the effects of other intrinsic and extrinsic factors on the regulation of enzymatic maturation in fetal small intestine.
Anat Rec 1984 Sep
PMID:Effect of dexamethasone on the fetal mouse small intestine in organ culture. 638 76

To establish the mode of fertilization in a marsupial, a morphological investigation was made of the gametes of the South American grey short-tailed opossum. Monodelphis domestica, at the time of fertilization in vivo and in vitro. Oestrus was induced in females by the introduction of an unfamiliar male. To obtain oocytes recently fertilized in vivo, females were killed 18-24 hours after the first mating and the region of the oviduct containing eggs excised and fixed. Unfertilized mature oocytes were recovered from ovarian follicles 15-18 hours after first mating and fertilized in vitro with cauda epididymal spermatozoa in a modified MEM medium supplemented with bovine serum albumin at 37 degrees C in 5% CO2 in air. Following sperm-egg binding and fertilization, oocytes were fixed and prepared for light and electron microscopy. Spermatozoa unpaired prior to fertilization in vivo and in vitro and single spermatozoa bound to the zona surface by their plasmalemma overlying the acrosome on the dorsal face of the sperm head. The acrosome reaction was only observed at the zona surface (suggesting that it may be induced by zona components) and involved a vesiculation of sperm plasma and acrosomal membranes over the main body of the acrosome but not over the narrow, marginal region which persisted after the acrosome reaction was complete. Sperm penetration of the zona pellucida caused a large breach in the zona and the dispersal of perivitelline material. The fusion of the spermatozoon with the oolemma occurred first over the marginal acrosomal region and was accompanied by a fertilization cone which protruded through the zona penetration hole. Activation of the egg was characterized by the release of material from vesicles in the peripheral cytoplasm and extrusion of the second polar body. The mode of fertilization in Monodelphis was compared with what is known in other marsupials (New World and Australian) and eutherian (placental) mammals. It was concluded that the general features of the acrosome reaction and sperm-egg fusion may be essentially similar in both groups and that an evolutionary schism did not occur following the development of the eutherian mode of fertilization.
Anat Rec 1993 Sep
PMID:Ultrastructural characteristics of in vivo and in vitro fertilization in the grey short-tailed opossum, Monodelphis domestica. 821 40

Alcaligenes eutrophus A5 catabolizes biphenyl to CO2 via benzoate and 4-chlorobiphenyl to 4-chlorobenzoate. In curing and conjugation experiments, the A5 endogenous 51-kb IncP1 plasmid pSS50 was found to be dispensable for biphenyl and 4-chlorobiphenyl catabolism. Transfer of the biphenyl- and 4-chlorobiphenyl-degrading phenotype by means of pSS50 was observed at a frequency of 10(-5) per transferred plasmid in matings of A5 with other A. eutrophus strains. Transconjugants harbor enlarged pSS50 derivatives which contain additional genetic information governing the oxidation of biphenyl and 4-chlorobiphenyl to benzoate and 4-chlorobenzoate and originating from the chromosome of strain A5. The following observations indicate that the catabolic genes reside on a 59-kb large transposon (Tn4371) for which a restriction map is presented. (i) Tn4371 transposes between different replicons and at different locations of the same replicon. (ii) Transposition was observed in a Rec- strain of A. eutrophus. (iii) Tn4371 transposes as a single, contiguous piece of DNA. Although an RP4::Tn4371 plasmid was stably maintained in different hosts, the plasmid conferred growth on biphenyl only when present in strains of A. eutrophus and in an Acinetobacter sp. strain.
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PMID:Identification of a catabolic transposon, Tn4371, carrying biphenyl and 4-chlorobiphenyl degradation genes in Alcaligenes eutrophus A5. 838 64

Previous work has shown that cell proliferation is a major contributor to the early palate morphogenesis in mammals. The present study was undertaken to examine the effect of EGF, TGFbeta1 and their combination on proliferation (measured by DNA synthesis) and on the expression of a growth related proto-oncogene, c-myc, in embryonic hamster palate mesenchymal cells (HPMC). Vertically developing hamster palatal shelves were dissected on day 11 of gestation, and trypsinized, and primary cultures were grown in DMEM + 10% serum at 37 degrees C and 5% CO2. Following appropriate growth factor treatment of HPMC, DNA synthesis was measured by scintillation counting and extracted RNA was subjected to Northern blot analysis. In serum-starved, pre-confuent cultures treated with EGF (20 ng/ml), DNA synthesis was stimulated in the presence of 2.5% serum. In contrast, treatment of HPMC with TGFbeta1 (10 ng/ml) in the presence or absence of EGF/serum for 24 hr, or HPMC pre-treatment with TGFbeta1 (30 min) followed by EGF/serum (24 hr), resulted in an arrest of DNA synthesis. Northern blot analysis of RNA extracted from HPMC showed that as serum-starved, growth-arrested cells progressed through G0 to G1 phase of the cell cycle, following EGF treatment, c-myc was expressed by 1 hr and declined thereafter. In contrast, TGFbeta1 did not support expression of c-myc. Following pre- or co-treatment with TGFbeta1, the EGF +/- serum-induced expression of c-myc was seen between 1 and 6 hr. It appears that EGF-induced expression of c-myc may be involved in advancing the HPMC in G1, and thus may contribute to the onset of DNA synthesis in HPMC. Since co- or pre-treatment with TGFbeta1 did not inhibit EGF/serum induced expression of c-myc, it is possible that growth arresting effect of TGFbeta1 may not be exerted directly through inhibition or blockage of c-myc expression.
Anat Rec 1999 04 01
PMID:Effects of EGF and TGFbeta1 on c-myc gene expression and DNA synthesis in embryonic hamster palate mesenchymal cells. 1020 53

Cellulose comprises a major portion of biomass on the earth, and the turnover of this material contributes to the CO2 cycle. Cellulases, which play a major role in the turnover of cellulosic materials, have been found either as free enzymes that work synergistically, or as an enzyme complex called the cellulosome. This review summarizes some of the general properties of cellulosomes, and more specifically, the properties of the Clostridium cellulovorans cellulosome. The C cellulovorans cellulosome is an extracellular enzyme complex with a molecular weight of about 1 x 10(6), and is comprised of at least ten subunits. The major subunit is the scaffolding protein CbpA, with a molecular weight of 189,000. This nonenzymatic subunit contains a cellulose binding domain (CBD) that binds the cellulosome to the substrate, nine conserved cohesins or enzyme binding domains, and four conserved surface layer homologous (SLH) domains. It is postulated that the SLH domains help to bind the cellulosome to the cell surface. The cellulosomal enzymes include cellulases (family 5 and 9 endoglucanases and a family 48 exoglucanase), a mannanase, a xylanase, and a pectate lyase. The cellulosome is capable of converting Arabidopsis and tobacco plant cells to protoplasts. One of the endoglucanases, EngE, contains three tandemly repeated SLHs at its N-terminus, and therefore appears capable of binding to the scaffolding protein CbpA as well as to the cell surface. Cellulosomes can attack crystalline cellulose, but the free cellulosomal enzymes can attack only soluble and amorphous celluloses. Nine genes for the cellulosome are found in a gene cluster cbpA-exgS-engH-engK-hbpA-engL-manA-engM-engN. Other cellulosomal genes such as engB, engE, and engY are not linked to the major gene cluster or to each other. By determining the structure and function of the cellulosome, we hope to increase the efficiency of the cellulosome by genetic engineering techniques.
Chem Rec 2001
PMID:The Clostridium cellulovorans cellulosome: an enzyme complex with plant cell wall degrading activity. 1189 54

In situ time-resolved Fourier transform infrared (FTIR) and microprobe Raman spectroscopies were used to characterize the reaction mechanisms of the partial oxidation of methane to syngas over SiO(2)- and gamma-Al(2)O(3)-supported rhodium and ruthenium catalysts. The interaction of both pure methane and a methane/oxygen mixture at a stoichiometric feed ratio with an oxygen-rich catalyst surface led to the formation of CO2 and H(2)O as the primary products. For the H(2)-pretreated samples, the reaction mechanisms with the catalysts differ. Only Rh/SiO(2) is capable of catalyzing the direct oxidation of methane to syngas, while syngas formation over Rh/gamma-Al(2)O(3), Ru/SiO(2), and Ru/gamma-Al(2)O(3) can be achieved mainly via a combustion-reforming scheme. The significant difference in the mechanisms for partial oxidation of methane to syngas over the catalysts can be correlated to the differences in the concentration of oxygen species (O(2-)) on the catalyst surface during the reaction, mainly due to the difference in the nature of the metals and supports.
Chem Rec 2002
PMID:Mechanistic study of partial oxidation of methane to syngas using in situ time-resolved FTIR and microprobe Raman spectroscopies. 1200 Dec 9

The reaction of M2(O2CBu(t))4 (M = Mo, W) with a dicarboxylic acid in toluene yields compounds of general formula [M2]-O2C-X-CO2-[M2] ([M2] = M2(O2CBu(t))3; X = conjugated spacer). The M2 units are electronically coupled via interactions between the M2 delta and dicarboxylate pi* orbitals, and the magnitude of this coupling is revealed by electronic structure calculations and spectroscopic data. These compounds show intense metal to ligand charge transfer (MLCT) absorptions in the visible region of the electronic spectrum that are temperature and solvent dependent. Evidence of electronic coupling is seen in their cyclic voltammograms, which show two successive one-electron oxidations. The extent of electronic coupling in the mixed valence radical cations [M2]-O2C-X-CO2-[M2]+, generated by oxidation with one equivalent of AgPF6 or FeCp2PF6, is evaluated by EPR and UV-vis-NIR spectroscopic data, and delocalized behavior is observed in compounds with W2 units separated by up to 13.6 angstroms. The simplicity of the frontier M2 orbital interactions with the bridge pi orbitals provides a convenient system with which to study electron transfer in mixed valence systems, as compared to the extensively studied, but more complicated, dinuclear t(2g)6/t(2g)5 mixed valence compounds. Oligomeric and polymeric compounds incorporating M2 units have also been synthesized, having general formula [M2(O2CR)2(O2C-Thio-CO2)]n (Thio = n-hexyl substituted ter- and quinque-thiophenes). They can be deposited as thin films by spin coating, and show photoluminescence and electroluminescence. These metallo-polythiophenes show potential for application in electronic materials. (
Chem Rec 2005
PMID:Electronically-coupled MM quadruply-bonded complexes of molybdenum and tungsten. 1621 87

Methanol is a valuable raw material used in the manufacture of useful chemicals as well as a potential source of energy to replace coal and petroleum. Biotechnological interest in the microbial utilization of methanol has increased because it is an ideal carbon source and can be produced from renewable biomass. Formaldehyde, a cytotoxic compound, is a central metabolic intermediate in methanol metabolism. Therefore, microorganisms utilizing methanol have adopted several metabolic strategies to cope with the toxicity of formaldehyde. Formaldehyde is initially detoxified through trapping by some cofactors, such as glutathione, mycothiol, tetrahydrofolate, and tetrahydromethanopterin, before being oxidized to CO2. Alternatively, free formaldehyde can be trapped by sugar phosphates as the first reaction in the C1 assimilation pathways: the xylulose monophosphate pathway for yeasts and the ribulose monophosphate (RuMP) pathway for bacteria. In yeasts, although formaldehyde generation and consumption takes place in the peroxisome, the cytosolic formaldehyde oxidation pathway also plays a role in formaldehyde detoxification as well as energy formation. The key enzymes of the RuMP pathway are found in a variety of microorganisms including bacteria and archaea. Regulation of the genes encoding these enzymes and their catalytic mechanisms depend on the physiological traits of these organisms during evolution.
Chem Rec 2005
PMID:Assimilation, dissimilation, and detoxification of formaldehyde, a central metabolic intermediate of methylotrophic metabolism. 1627 35

Proton dissociation of an aqua-Ru-quinone complex, [Ru(trpy)(q)(OH2)]2+ (trpy = 2,2' : 6',2''-terpyridine, q = 3,5-di-t-butylquinone) proceeded in two steps (pK(a) = 5.5 and ca. 10.5). The first step simply produced [Ru(trpy)(q)(OH)]+, while the second one gave an unusual oxyl radical complex, [Ru(trpy)(sq)(O-*)]0 (sq = 3,5-di-t-butylsemiquinone), owing to an intramolecular electron transfer from the resultant O2- to q. A dinuclear Ru complex bridged by an anthracene framework, [Ru2(btpyan)(q)2(OH)2]2+ (btpyan = 1,8-bis(2,2'-terpyridyl)anthracene), was prepared to place two Ru(trpy)(q)(OH) groups at a close distance. Deprotonation of the two hydroxy protons of [Ru2(btpyan)(q)2(OH)2]2+ generated two oxyl radical Ru-O-* groups, which worked as a precursor for O2 evolution in the oxidation of water. The [Ru2(btpyan)(q)2(OH)2](SbF6)2 modified ITO electrode effectively catalyzed four-electron oxidation of water to evolve O2 (TON = 33500) under electrolysis at +1.70 V in H2O (pH 4.0). Various physical measurements and DFT calculations indicated that a radical coupling between two Ru(sq)(O-*) groups forms a (cat)Ru-O-O-Ru(sq) (cat = 3,5-di-t-butylcathechol) framework with a mu-superoxo bond. Successive removal of four electrons from the cat, sq, and superoxo groups of [Ru2(btpyan)(cat)(sq)(mu-O2-)]0 assisted with an attack of two water (or OH-) to Ru centers, which causes smooth O2 evolution with regeneration of [Ru2(btpyan)(q)2(OH)2]2+. Deprotonation of an Ru-quinone-ammonia complex also gave the corresponding Ru-semiquinone-aminyl radical. The oxidized form of the latter showed a high catalytic activity towards the oxidation of methanol in the presence of base. Three complexes, [Ru(bpy)2(CO)2]2+, [Ru(bpy)2(CO)(C(O)OH)]+, and [Ru(bpy)2(CO)(CO2)]0 exist as an equilibrium mixture in water. Treatment of [Ru(bpy)2(CO)2]2+ with BH4- gave [Ru(bpy)2(CO)(C(O)H)]+, [Ru(bpy)2(CO)(CH2OH)]+, and [Ru(bpy)2(CO)(OH2)]2+ with generation of CH3OH in aqueous conditions. Based on these results, a reasonable catalytic pathway from CO2 to CH3OH in electro- and photochemical CO2 reduction is proposed. A new pbn (pbn = 2-pyridylbenzo[b]-1,5-naphthyridine) ligand was designed as a renewable hydride donor for the six-electron reduction of CO2. A series of [Ru(bpy)(3-n)(pbn)n]2+ (n = 1, 2, 3) complexes undergoes photochemical two- (n = 1), four- (n = 2), and six-electron reductions (n = 3) under irradiation of visible light in the presence of N(CH2CH2OH)3.
Chem Rec 2009
PMID:Metal-catalyzed reversible conversion between chemical and electrical energy designed towards a sustainable society. 1950 3

Postnatal mixed respiratory-metabolic acidosis is common in calves, and depending on its severity can impair vitality or even cause death. Carbon dioxide accounts for the respiratory component and L-lactate for the metabolic component of the mixed acidosis, but it remains unclear which component determines the severity and duration of the acidosis. In a first attempt to clarify, this was investigated retrospectively in 31 calves during the first two hours of life, and in 13 calves during the first three days of life. Venous blood was collected for blood gas analysis and measurement of acid-base variables and L-lactate concentration. pH Was more strongly correlated with L-lactate concentration (r(2)=0.808) than with partial pressure of CO2 (pCO2, r(2)=0.418). Duration of parturition had a distinct effect on pH and L-lactate concentration but not on pCO2; calves born within six hours of rupture of the allantoic sac had a higher pH and lower L-lactate concentration than calves born after a longer duration of parturition (both P<0.01). Normalisation of pCO2 took four hours and normalisation of L-lactate took 48 hours. It was concluded that L-lactate is a more important factor in the pathogenesis of acidosis than pCO2, and that the duration of metabolic acidosis exceeds that of respiratory acidosis in perinatal asphyxia of calves.
Vet Rec 2013 May 18
PMID:The effect of lactic acidosis on the generation and compensation of mixed respiratory-metabolic acidosis in neonatal calves. 2348 6


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