Spontaneous Omental Infarction: An Unusual Etiology of Abdominal Pain.

A 49-year-old lady with previous scars complained of acute abdominal pain for two days. Her right hypochondrium was tender and guarding upon assessment. The laboratory investigations were unremarkable. Due to a diagnostic incongruity, computed tomography of the abdomen was performed showing a suspicious lesion at anterolateral aspect of the ascending colon. Surgical intervention was decided and intraoperative finding was consistent with spontaneous omental infarction. Omentectomy was undertaken and final histology was compatible with the intraoperative diagnosis. Although it is exceptional, omental infarction should be considered as part of the differential diagnoses of right-sided acute abdominal pain with normal laboratory investigations. This case highlights its unexpected discovery and we describe its literature reviews.

COMMUNITY CO-DESIGNED SCHISTOSOMIASIS CONTROL INTERVENTIONS FOR SCHOOL-AGED CHILDREN IN ZANZIBAR.

Top-down biomedical interventions to control schistosomiasis in sub-Saharan Africa have had limited success, primarily because they fail to engage with the social, political, economic and ecological contexts in which they are delivered. Despite the call to foster community engagement and to adapt interventions to local circumstances, programmes have rarely embraced such an approach. This article outlines a community co-designed process, based upon Human-Centered Design, to demonstrate how this approach works in practice. It is based on initial work undertaken by social science researchers, public health practitioners and community members from the Zanzibar Islands, Tanzania, between November 2011 and December 2013. During the process, 32 community members participated in a qualitative and quantitative data-driven workshop where they interpreted data on local infections from S. haematobium and co-designed interventions with the assistance of a facilitator trained in the social sciences. These interventions included the implementation of novel school-based education and training, the identification of relevant safe play activities and events at local schools, the installation of community-designed urinals for boys and girls and the installation of community-designed laundry-washing platforms to reduce exposure to cercariae-contaminated fresh water. It is suggested that the a community co-designed process, drawing from Human-Centered Design principles and techniques, enables the development of more sustainable and effective interventions for the control of schistosomiasis.

Structural heterogeneity of Pseudomonas aeruginosa bacterioferritin.

The subunit composition, amino acid sequence and haem-binding characteristics of bacterioferritin (BFR) from Pseudomonas aeruginosa have been studied. Unlike other BFRs, P. aeruginosa BFR was found to contain two subunit types, designated alpha and beta, which differed considerably in their amino acid sequences. The N-terminal 69 and 55 amino acids of the alpha and beta subunits respectively were determined. The alpha subunit differed most from other BFRs. The two subunits were present in variable proportions in different preparations. The maximum stoichiometry of haem binding was found to be sample-dependent and to be different from the previously reported one per subunit [Kadir and Moore (1990) FEBS Lett. 271, 141-143]. This previous haem-binding study was shown to have been carried out with damaged protein, which contained both normal alpha and beta subunits and shorter versions of these that appeared to have been produced by cleavage of the normal subunits. The possibility that aging processes degrade ferritins and affect their haem-binding characteristics is discussed.

Haem and non-haem iron sites in Escherichia coli bacterioferritin: spectroscopic and model building studies.

The bacterioferritin (BFR) of Escherichia coli is an iron-storage protein containing 24 identical subunits and between three and 11 protohaem IX groups per molecule. Titration with additional haem gave a maximum loading of 12-14 haems per molecule. The e.p.r. spectra and magnetic c.d. spectra of the protein-bound haem show it to be low-spin Fe(III), and coordinated by two methionine residues as previously reported for BFRs isolated from Pseudomonas aeruginosa and Azotobacter vinelandii [Cheesman, Thomson, Greenwood, Moore and Kadir, Nature (London) (1990) 346, 771-773]. A recent sequence alignment indicated that BFR may be structurally related to ferritin. The molecular model proposed for E. coli BFR has a four-alpha-helix-bundle subunit conformation and a quaternary structure similar to those of mammalian ferritins. In this model there are two types of hydrophobic pocket within which two methionine residues are correctly disposed to bind haem. The e.p.r. spectra also reveal a monomeric non-haem Fe(III) species with spin, S = 5/2. On the basis of sequence comparisons, a ferroxidase centre has recently been proposed to be present in BFR [Andrews, Smith, Yewdall, Guest and Harrison (1991) FEBS Lett. 293, 164-168] and the possibility that this Fe(III) ion may reside at or near the ferroxidase centre is discussed.

Tissue reaction after intramuscular injection of liposomes in mice.

Liposomes are effective carrier systems for prolonged drug release. As all other drug formulations for parenteral use, the safety of liposomal formulations should be established before clinical application. In this study, some safety aspects of intramuscularly injected (single dose) "gel-state" type liposomes and the ability of liposome encapsulation to diminish irritating effects of intramuscularly applied drugs were studied by histopathological analysis over a period of 14 days in mice. Injection of saline solution showed no tissue reaction at the injection site. Intramuscular injection of liposomes alone showed an infiltrative reaction consisting of a population of macrophages. Within this population fat cells were present. In time, the population of macrophages present at the injection site was largely replaced by loose connective tissue. Novaminsulfon (NS) injected intramuscularly in "free" form is a strongly irritating drug, causing hemorrhage, cell necrosis, inflammatory reactions and eventually fibrosis. However, NS being encapsulated in liposomes was hardly more irritating than liposomes alone. The same was true for liposome-encapsulated chloroquine and free chloroquine. When sustained-release of a drug is therapeutically desirable, the parenteral application of a liposome-encapsulated formulation can be considered for drugs, in particular for those drugs causing tissue injury at the injection site.

Spectroscopic identification of the haem axial ligands of haemoferritin and location of possible haem-binding sites in ferritin by molecular modelling.

Horse spleen ferritin will bind up to 16 protoporphyrin IX haem groups per 24 subunits in vitro [Kadir & Moore (1990) FEBS Lett. 276, 81-84] at a site that causes the haem to be low spin for both ferric and ferrous states. E.p.r. spectra at 10 K of the oxidized form of the resulting haemoferritin gives g values of 2.93, 2.26 and 1.55, characteristic of low-spin haem. The near-i.r. magnetic circular dichroism spectrum shows a porphyrin-to-ferric charge-transfer band at 1590 nm. The spectroscopic parameters indicate that the haem group is probably bound by two histidine ligands. Molecular modelling studies reveal one type of potential haem-binding site in horse L-chain ferritin with bis-histidine co-ordination. This is an intersubunit site which lies in a pocket within the ferritin protein shell in the region of the 3-fold channel. The ligands are His-114 and His-124 in horse L-chain. A second possible set of sites in human H-chain ferritin involves His-60 residues in the pockets between pairs of subunits. These are considered less likely sites of haem occupancy. There are three of the intersubunit sites in horse L-chain ferritin at each of the eight 3-fold channels. We propose that conformational crowding between haem-binding sites at a given channel prevents more than two haems per channel being bound.

E.p.r. and magnetic circular dichroism spectroscopic characterization of bacterioferritin from Pseudomonas aeruginosa and Azotobacter vinelandii.

The e.p.r. and magnetic circular dichroism (m.c.d.) spectra of bacterioferritin (BFR) extracted from Pseudomonas aeruginosa and Azotobacter vinelandii have been studied over a wide temperature range down to liquid-helium temperature. The e.p.r. spectra show the presence of low-spin Fe3+ haem with g values of 2.86, 2.32, 1.48 (P. aeruginosa) and 2.88, 2.31, 1.46 (A. vinelandii), in both the presence and absence of the BFR core. Together with evidence from the porphyrin-to-Fe3+ charge-transfer band at 2240 and 2270 nm the axial haem ligands are identified as two methionines. The low-temperature m.c.d. spectra in the region 300-1000 nm of P. aeruginosa and A. vinelandii BFR are identical with one another and unaffected by removal of the iron core. Hence it can be concluded that the presence of the iron core has no detectable effect on the electronic states and on the stereochemistry of the haem group. This was unexpected, in view of the observations by Watt, Frankel, Papaefthymiou, Spartalian & Stiefel [(1986) Biochemistry 25, 4330-4336] that the redox potential of the haem group in A. vinelandii BFR shifts from -475 mV to -225 mV on removal of the core. The e.p.r. spectra of holoBFR show a broad symmetrical derivative-shaped band centred at g = 2.0 which decreases in bandwidth as the temperature is raised. This signal is assigned to the uncompensated electron spins of the iron core.

Haem binding to ferritin and possible mechanisms of physiological iron uptake and release by ferritin.

Haem binding to horse spleen ferritin and Pseudomonas aeruginosa bacterioferritin has been studied by spectroscopic methods. A maximum of 16 haems per ferritin molecule, and 24 haems per bacterioferritin molecule, has been shown to bind. The influence of the bound haem on the rate of reductive iron release has been investigated. With a range of reductants and in the absence of haem the rate of release varied with the reductant, but in the presence of haem the rate was both independent of the reductant and faster than with any of the reductants alone. This indicates the rate-limiting step for iron release in the absence of haem was electron-transfer across the protein shell. Based on the results obtained with the in vitro assay system and from a consideration of data currently in the literature, plausible schemes for ferritin and bacterioferritin iron uptake and release are described.

Spectroscopic studies of Rhodobacter capsulatus cytochrome c' in the isolated state and in intact cells.

Ferricytochrome c' from Rhodobacter capsulatus was investigated by 1H-NMR, EPR and optical spectroscopies. A haem-linked ionisation, occurring with a pKa of 8.4 at 25 degrees C, was observed and assigned to the ionisation of the axial histidine ligand by comparison with data for related proteins. At pH values below this pKa the spin-state of the haem Fe3+ is shown to be a quantum mechanically admixed S = 3/2, 5/2 state. Above the pKa the Fe3+ is high-spin. EPR studies of intact cells grown photoheterotrophically reveal that in situ cytochrome c' exists largely in the ferrous state. Upon the addition of [Fe(CN)6]3- the protein becomes oxidised and EPR spectra reveal that the Fe3+ spin-state is a quantum mechanically admixed S = 3/2, 5/2 state. These data indicate that the unusual spin-state of ferricytochrome c' is not a consequence of changes to the protein on its isolation, as had been suggested previously. They also indicate that in situ cytochrome c' is located in an environment with a pH less than 7.

Animal ferritin and bacterioferritin contain quinones.

The origin of the 440 nm fluorescence of horse spleen ferritin and of Pseudomonas aeruginosa and Azotobacter vinelandii bacterioferritin has been investigated using a Nitro Blue Tetrazolium/glycinate colorimetric test specific for quiones [Paz, Flückiger, Boak, Kagan & Gallop (1991) J. Biol. Chem. 266, 689-692]. The results of the analysis indicate that ferritin and bacterioferritins contain quinones. A possible functional role of these quinones in iron uptake and release is described, as is the possibility that the presence of quinones in these proteins results from oxidative damage.

Haem binding to horse spleen ferritin and its effect on the rate of iron release.

Horse spleen ferritin is shown to bind haem to generate a haemoprotein, named herein haemoferritin. A total of 14-16 haem molecules are bound per 24 subunits of ferritin. The molecular mass of the non-haem-iron-free haemoferritin has been determined to be 420 +/- 40 kDa, indicating that haem binding does not lead to dissociation of the 24 subunits that comprise the ferritin molecule. The functional role of the bound haem has been investigated with respect to the release of iron from the non-haem iron core. The bound haem is shown to increase the rate of iron release in a reductive assay system. In the absence of haem the rate of iron release depends on the redox potential of the reductant, but in the presence of haem the rate is largely independent of the reductant and is faster than the rate for the haem-free ferritin. These data haem, but in the presence of haem electron transfer is not rate-limiting.

Aluminium transport in blood serum. Binding of aluminium by human transferrin in the presence of human albumin and citrate.

The binding of Al3+ by human serum transferrin has been investigated by u.v.-visible difference spectroscopy. In the presence of 25 mM-HCO3- at pH 7.4, the apparent association constants were found to be 1.69 x 10(12) M-1 and 5.36 x 10(11) M-1. These association constants are pH-dependent, reducing with both increasing and decreasing pH. The apparent pKa values were found to be 6.7 and 8.2. Competitive assays of binding of Al3+ to transferrin in the presence of citrate and human serum albumin at molar ratios corresponding to those found in normal plasma showed that a considerable amount of Al3+ was not bound to transferrin. Taking a concentration of 5 microM as a typical value observed for the plasma of patients on haemodialysis [Harris & Sheldon (1990) Inorg. Chem. 29, 119-124] the competitive binding assay indicate that approximately 60% of it is bound to transferrin, approximately 34% to albumin and the remainder to citrate. These results therefore suggest that, although transferrin at pH 7.4 is the major Al(3+)-binding component of plasma, an appreciable amount of Al3+ present in patients on haemodialysis may be bound to albumin.

Mössbauer spectroscopic studies of iron in Pseudomonas aeruginosa.

The present Mössbauer spectroscopic studies of isolated bacterioferritin and whole cells of Pseudomonas aeruginosa have shown that the iron core of bacterioferritin is not altered on isolation. These studies have also shown that the bacterioferritin core is typically 85% oxidized within the cell and may contain a significant proportion of its iron as small clusters during the early stage of the stationary phase of cell growth.

Electron transfer between horse ferritin and ferrihaemoproteins.

Reactions of reduced horse spleen ferritin with horse and Saccharomyces cerevisiae ferricytochromes c, cow ferricytochrome b5, sperm-whale metmyoglobin and Pseudomonas aeruginosa ferricytochrome c-551 were investigated by u.v.-visible spectrophotometry. In all cases the reduced ferritin reduced the ferrihaemoproteins. The rate of reduction varied from less than 0.2 M-1.s-1 for metmyoglobin to 1.1 x 10(3) M-1.s-1 for horse ferricytochrome c (0.1 M-phosphate buffer, pH 7.4, at 25 degrees C). We conclude that the mechanism of ferrihaemoprotein reduction involves long-range electron transfer through the coat of ferritin and that such electron transfer is rapid enough to account for the rates of iron release observed by other workers in reductive release assays.

The role of drug lipophilicity in release from intra-adipose and intramuscular injection sites.

Release rates from intramuscular and intra-adipose injection sites are dependent upon several including injection depth. Little is known about the relationship between drug lipophilicity and transport rate of drugs through adipose and muscle tissue. The principal objective of the present study was to investigate to what extend drug lipophilicity affects release and release rate from adipose tissue. Nine pigs were given intravenous (0.1 mg/kg), intramuscular (0.2 mg/kg) and intra-adipose (0.2 mg/kg) injections of propranolol, alprenolol, carazolol, metoprolol and atenolol. The fraction not-absorbed versus time plots after intramuscular and intra-adipose injection showed a biphasic decline for all model compound with the exception of atenolol being the most hydrophilic drug. This biphasic decline indicates that two different mechanisms may be involved in drug release. Initial release rates after intra-adipose injection were negatively correlated (Kendall's rankorder test) with fat-buffer distribution constants. The extent of release after 24 h could be considered as a characteristic parameter for the second release phase. The amounts released after 24 h were lower for propranolol, alprenolol, carazolol and metoprolol than for atenolol. Incomplete release at 24 h can be explained by the decrease of the solvent drag after absorption of the vehicle is complete.

Haem binding to horse spleen ferritin.

Horse spleen ferritin, a spherical protein shell of 24 subunits, contains no haem when extracted. This contrasts with ferritins isolated from bacterial sources which have the capacity to bind up to 24 haem groups [(1990) FEBS Lett. 271, 141-143] via two methionine residues [(1990) Nature 341, 771]. Here it is shown that horse spleen ferritin can bind between 15 and 17 haems per 24 subunits with an apparent association constant of 2.2-3.2 x 10(4) M-1. The strength of haem binding appears to be unaffected either by the presence of the core or by the oxidation state of the haem. The demonstration of the ability of animal ferritin to bind haem strengthens the similarity between it and bacterioferritin and could have major consequences for its mechanism of action in physiological iron uptake and release processes.

Pharmacokinetics of intravenously, intramuscularly and intra-adiposely administered carazolol in pigs.

The beta-blocking agent carazolol is used for the prevention of stress syndromes in pigs. Little is known of the pharmacokinetics of this drug, and therefore of its residue status in meat. In this study carazolol pharmacokinetics were investigated in a randomized three-way cross-over design in five pigs. A dose of 0.025 mg/kg was given intravenously, intramuscularly and intra-adiposely (in the subcutaneous fat layer). Carazolol was rapidly distributed and had a short half-life of 1.2-4.2 h. The distribution volume was calculated to be 0.22-0.65 l/kg. After intramuscular or intra-adipose administration the absorption pattern was biphasic. A rapid initial phase was followed by a slow second phase. Absorption was found to be incomplete at 24 h after intramuscular and intra-adipose administration ranging from 24 to 59% and 25 to 66%, respectively. The biphasic behaviour could be explained by retention of the drug in the tissues after absorption of the solvent was complete. A few hours after intravenous administration only negligible amounts of the drug were circulating in the body; however, considerable amounts of drug might have remained at the intramuscular or intra-adipose injection site.

Evidence for Mössbauer spectroscopy for different forms of iron core in Pseudomonas aeruginosa bacterial ferritin.

Mössbauer spectroscopic studies of whole cells of Pseudomonas aeruginosa, grown under different conditions, indicate that the predominant form of iron in the cells varies significantly. These differences are interpreted in terms of differences in the nature of the iron cores of the bacterial ferritin, which result from different growth conditions.

Bacterial ferritin contains 24 haem groups.

Pseudomonas aeruginosa bacterioferritin, also known as cytochrome b1 or cytochrome b557, has been isolated with 9 haems per 24 subunits. Various forms of the protein have been prepared including the completely haem-free protein and the fully haem-loaded protein with 24 haems per 24 subunits. The presence of the core does not significantly affect haem addition or removal. The absorbance ratio of the non-haem-iron-loaded protein, 278 nm:417 nm (oxidised), can be used to estimate the haem loading.

Bis-methionine axial ligation of haem in bacterioferritin from Pseudomonas aeruginosa.

The iron-containing bacterioferritins contain the protoporphyrin IX haem group. It has been established that Escherichia coli cytochrome b1, cytochrome b557 and bacterioferritin are identical. The optical spectra at room temperature of the haem group show it to be predominantly low-spin in both the ferrous and ferric states. The nature of the axial ligands binding the haem group to the polypeptide has, however, remained unknown. Low-spin, bis-coordinate haem centres in proteins typically have a role in rapid electron transfer as redox changes at the metal ion lead to little structural rearrangement. There are only four amino acids with side-chains that have ligand field strengths sufficient to generate the low-spin state of haem, namely, histidine, lysine, methionine and cysteine. Hence there are, potentially, ten different pairs of these four ligands which could be discovered in electron transfer haemoproteins. To date only three have been established with certainty. They are bis-histidine, as in mammalian cytochrome b5, methionine-histidine, typified by cytochrome c and lysine-histidine, recently recognized by spectroscopic methods in cytochrome f. Here we report the electron paramagnetic resonance and near infrared magnetic circular dichroism spectra of the oxidized state of Ps. aeruginosa bacterioferritin which enable the axial ligands to be identified as the thioether side chains of two methionine residues, a ligation scheme not previously reported for haem in any protein.