Biomedical subjects

M J Miller

Publications and source records attributed to M J Miller.

At least 181 records · Page 10Linked to original sources

PubMed1996

Variable responses of species and strains to white mineral oils and paraffin waxes.

Recent dietary studies on mineral hydrocarbon (MHC) white oils and waxes have shown inflammatory effects in Fischer 344 (F-344) rats, but not in other rat strains or dogs. Histopathologic effects seen in F-344 rats include mesenteric lymph node histiocytosis, liver granulomas, and inflammation of the mitral valve (only seen with paraffin waxes). Human ingestion of MHC can result in noninflammatory lipogranulomas (oil droplets) in tissues which are regarded as clinically unimportant. It is speculated that inherent interspecies differences in pharmacokinetics and/or immune function may contribute to the differential response to MHC seen in F-344 rats. The F-344 rat retains greater amounts of MHC in target tissues compared to other rat strains and dogs and appears to be more sensitive immunologically to MHC than other species, including humans. This strain may be predisposed to these effects as indicated by a high background incidence of inflammatory granulomatous lesions in control female F-344 rats. Because of its apparently unique sensitivity, relevance of effects seen in F-344 rat to human health is questionable and requires further investigation.

Animals↗

PubMed1996

Synthesis of optically pure chrysobactin and immunoassay development.

Chrysobactin (alpha-N-(2,3-dihydroxybenzoyl)-D-lysyl-L-serine), a siderophore that is essential for systemic virulence by plant pathogenic Erwinia chrysanthemi, was synthesized with high diastereomeric purity. Chrysobactin was prepared by coupling the N-hydroxysuccinimide ester of alpha-N-(2,3-dibenzyloxybenzoyl)-epsilon-N-Cbz-D-lysine with L-serine benzyl ester followed by deprotection via hydrogenolysis. Optically pure chrysobactin was obtained with 98% overall yield. A monoclonal antibody to ferric chrysobactin was developed and characterized as IgM. The antibody reacts with chrysobactin, ferric chrysobactin and less strongly with ferric dihydroxybenzoic acid. The antibody reacts weakly with the siderophores ferrichrome, A, ferric pseudobactin and ferric rhodotorulic acid. This antibody was used in a competitive immunoassay to detect ferric chrysobactin at 10(-8) to 10(-10) mol. This immunoassay may provide a useful method for the detection of chrysobactin in plant samples.

Animals↗

PubMed1996

Purification, spectroscopic analysis and biological activity of the macrocyclic dihydroxamate siderophore alcaligin produced by Bordetella pertussis and Bordetella bronchiseptica.

Hydroxamate siderophores of virulent Bordetella pertussis and Bordetella bronchiseptica strains were purified using a simple large-scale isolation procedure, and identified by various spectroscopic techniques as the macrocyclic dihydroxamate siderophore trivially known as alcaligin, 1,8(S),11,18(S)- tetrahydroxy-1,6,11,16-tetraazacycloeicosane-2,5,12,15-tetrone+ ++, which was previously isolated from the taxonomically-related bacterial species Alcaligenes denitrificans subsp. xylosoxydans. Alcaligin purified from iron-depleted cultures of B. pertussis and B. bronchiseptica exhibited specific growth-promoting activity under iron-restricted conditions for Bordetella indicator strains, and ere active in [55Fe]ferric alcaligin transport assays. Evidence suggests that several C2-symmetric conformations of alcaligin exist simultaneously in both methanolic and aqueous solution.

Alcaligenes↗

PubMed1996

Failure of L-NAME to cause inhibition of nitric oxide synthesis: role of inducible nitric oxide synthase.

We addressed the hypothesis that administration of nitric oxide synthase inhibitor, NG -nitro-L-arginine methyl ester (L-NAME) does not result in a sustained suppression of nitric oxide (NO) synthesis, because of a compensatory expression of inducible nitric oxide synthase (iNOS). L-NAME was administered in the drinking water (0.1-1.0 mg/ml) for 7 days to guinea pigs and rats. Nitric oxide synthesis was assessed by [1] ex vivo formation of nitrite in blood vessels and intestine [2] tissue levels of cGMP [3] iNOS gene expression by RT-PCR [4] NADPH diaphorase staining [5] direct assessment of NO release in tissue explants using a microelectrode/electrochemical detection system. Chronic L-NAME administration elevated intestinal cGMP and nitrite levels in guinea pigs (p < 0.05). In rats, intestinal nitrite levels were comparable in control and L-NAME treatment groups, whereas direct assessment of NO release defined a marked increase in the L-NAME group. Chronic L-NAME resulted in an induction of iNOS gene expression in rats and guinea pigs and novel sites of NADPH diaphorase staining in the intestine. We conclude that iNOS expression is responsible for a compensatory increase or normalization of NO synthesis during sustained administration of L-NAME.

Administration, Oral↗

PubMed1996

Biochemical characterization of mouse liver IBE1-amide immunoreactivity: limitations of antibody-based approaches for verification of peptide expression.

A high titer, specific antiserum, raised against a synthetic analogue of a unique peptide region within the human IGF-IB prohormone, detected specific immunoreactivity in extracts of mouse, chicken, sheep, and human liver. Specificity was confirmed by the ablation of immunoreactivity in the presence of excess synthetic immunogen. Here we report the isolation and characterization of one of the immunoreactive species from an extract of mouse liver: amino acid sequencing revealed that the purified product was 78% identical to the NH2-terminus of the alpha-subunit of mouse hemoglobin. Immunoblot analysis of a commercial preparation of mouse hemoglobin confirmed that the antiserum recognized hemoglobin. Addition of excess synthetic peptide to the antiserum eliminated the immunobinding to hemoglobin. The apparent "specificity" of even affinity-purified antiserum for hemoglobin provides a cautionary note for the interpretation of studies concluding antigen expression based solely on the presence of positive immunoreactivity.

Amino Acid Sequence↗

PubMed1996

Fetal growth retardation in rats may result from apoptosis: role of peroxynitrite.

Administration of the nitric oxide synthase (NOS) inhibitor NG-nitro-L-arginine methyl ester (L-NAME) results in fetal growth retardation. This study was designed to further examine the influence of NO on fetal growth, specifically, the potential role of inducible NOS and to evaluate the possibility that apoptosis contributed to uteroplacental dysfunction. L-NAME administration caused a paradoxical increase in NO synthesis determined by direct detection of NO by electrochemistry, nitrite accumulation, and cGMP levels, indicating that a lack of NO was not the cause of the fetal growth retardation. Additionally, supplemental L-arginine or NO donors failed to reverse the effects of L-NAME on fetal and placental size. Administration of low dose endotoxin (30 micrograms/kg IP daily for 6 d) also caused significant reductions in fetal and placental size and increased NO synthesis comparable to that seen with L-NAME. Inducible NOS was constitutively expressed in the pregnant uterus (smooth muscle and epithelia) and placenta (sinusoids and macrophages) but was absent in the nonpregnant state as determined by RT-PCR and immunohistochemistry. Neither L-NAME nor endotoxin modified the expression of iNOS. In situ evidence for apoptosis (DNA fragmentation) was minimal to absent in control pregnant rats, but markedly evident in the placenta (decidua) and uterus of rats treated with L-NAME or endotoxin. Immunohistochemical evidence for nitrotyrosine, a marker for peroxynitrite formation, was absent in control rats but colocalized with apoptosis in the L-NAME and LPS groups. We conclude that L-NAME-induced fetal growth retardation is not due to a lack of NO, but as for endotoxin, results from a net reduction in cellular proliferation due to the induction of apoptosis, possibly in response to peroxynitrite formation.

Animals↗

PubMed1996

Synthesis and in vitro antibacterial activity of spermidine-based mixed catechol- and hydroxamate-containing siderophore--vancomycin conjugates.

The first antibiotic conjugates of vancomycin (1) and siderophore analogues containing spermidine-based catechol ligands (conjugate 11) as well as mixed catechol and hydroxamate ligands (conjugate 13) are described. The design of the conjugates was based on the earlier observation that conjugation of siderophore components to beta-lactam antibiotics induced active iron transport-mediated drug delivery. The novel conjugates (11 and 13) were synthesized by selective acylation of the primary amino group of 1. Preliminary biological studies indicated that siderophore modified vancomycins lost some activity (4- to 16-fold) against Gram-positive bacteria relative to vancomycin itself, and were generally similar to vancomycin in activity against Gram-negative bacteria under iron-sufficient conditions. However, under iron-depleted conditions which mimic human serum, conjugate 11 displayed enhanced antibacterial activity against an antibiotic hypersensitive strain of Pseudomonas aeruginosa.

Anti-Bacterial Agents↗

PubMed1996

Detection of adrenomedullin, a hypotensive peptide, in amniotic fluid and fetal membranes.

OBJECTIVE: Our purpose was to determine whether adrenomedullin, a multifunctional regulatory peptide involved in blood flow regulation and growth stimulation and with antimicrobial activity, was a component of amniotic fluid from second-trimester human fetus and to determine the source of this peptide. STUDY DESIGN: A prospective descriptive study was performed on 134 patients undergoing amniocentesis after genetic counseling, ultrasonography, and informed consent. Adrenomedullin expression was determined by immunocytochemical analysis, Western blot analysis, reverse transcriptase-polymerase chain reaction, and in situ reverse transcriptase-polymerase chain reaction in fetal membranes and with radioimmunoassay in amniotic fluids. RESULTS: Radioimmunoassay of the 134 amniotic fluid specimens revealed adrenomedullin-like immunoreactivity in all of them, ranging in concentration from 10 to 300 fmol/25 microliters (170 +/- 62 fmol/25 microliters). Immunocytochemical analysis, Western blot analysis, reverse transcriptase-polymerase chain reaction, and in situ reverse transcriptase-polymerase chain reaction further established the expression of adrenomedullin protein and messenger ribonucleic acid in fetal amniotic membranes, suggesting that this organ is the source of amniotic adrenomedullin. CONCLUSIONS: Our results clearly demonstrate the presence of adrenomedullin in second-trimester human amniotic fluid and adrenomedullin messenger ribonucleic acid and protein in amniotic membranes, suggesting that adrenomedullin is a hormone involved in the maintenance of normal pregnancy. Further studies with these molecular tools are in progress to determine the precise role of this hormone and whether adrenomedullin plays a role in the pathogenesis of various disorders of pregnancy.

Adrenomedullin↗

PubMed1996

Iron transport-mediated drug delivery using mixed-ligand siderophore-beta-lactam conjugates.

BACKGROUND: Assimilation of iron is essential for microbial growth. Most microbes synthesize and excrete low molecular weight iron chelators called siderophores to sequester and deliver iron by active transport processes. Specific outer membrane proteins recognize, bind and initiate transport of species-selective ferric siderophore complexes. Organisms most often have specific receptors for multiple types of siderophores, presumably to ensure adequate acquisition of the iron that is essential for their growth. Conjugation of drugs to synthetic hydroxamate or catechol siderophore components can facilitate active iron-transport-mediated drug delivery. While resistance to the siderophore-drug conjugates frequently occurs by selection of mutants deficient in the corresponding siderophore-selective outer membrane receptor, the mutants are less able to survive under iron-deficient conditions and in vivo. We anticipated that synthesis of mixed ligand siderophore-drug conjugates would allow active drug delivery by multiple iron receptor recognition and transport processes, further reducing the likelihood that resistant mutants would be viable. RESULTS: Mixed ligand siderophore-drug conjugates were synthesized by combining hydroxamate and catechol components in a single compound that could chelate iron, and that also contained a covalent linkage to carbacephalosporins, as representative drugs. The new conjugates appear to be assimilated by multiple active iron-transport processes both in wild type microbes and in selected mutants that are deficient in some outer membrane iron-transport receptors. CONCLUSIONS: The concept of active iron-transport-mediated drug delivery can now be extended to drug conjugates that can enter the cell through multiple outer membrane receptors. Mutants that are resistant to such conjugates should be severely impaired in iron uptake, and therefore particularly prone to iron starvation.

Anti-Bacterial Agents↗

PubMed1996

Association of signaling proteins with a nonmitogenic heterodimeric complex composed of epidermal growth factor receptor and kinase-inactive p185c-neu.

The functional consequences of heterodimer formation between the epidermal growth factor receptor (EGFr) and the p185c-neu receptor tyrosine kinase include increased mitogenic and transformation potencies. To determine the possible alteration of signal transduction pathways resulting from this heteromeric complex, the capacity of several signaling proteins to associate with the heterodimeric receptors has been assayed. The in vivo interaction with the EGFr/p185c-neu heterodimer of several signal transduction proteins, including phospholipase C-gamma 1 (PLC-gamma 1), the p85 subunit of phosphotidylinositol 3-kinase, the ras GTPase activating protein, SHC, NCK, p72RAF, and the tyrosine phosphatase SHPTP2, was measured by coimmunoprecipitation. The binding of these signaling proteins to a complex composed of EGFr and a kinase-inactive form of p185 (p185K757M) was not impaired, even though the mitogenic and transformation activity of this complex had been abrogated. In addition, the EGF-induced phosphorylation of GAP, p85, and PLC-gamma 1 did not correlate with the dominant-negative action of p185K757M on EGFr function. Thus, substrate association and phosphorylation do not correlate stringently with the mitogenic and transforming activity of this receptor complex, suggesting additional pathways or mechanisms vital to EGFr/p185c-neu heterodimeric signaling.

Animals↗

PubMed1996

Aesthetic breast reconstruction using a combination of free transverse rectus abdominis musculocutaneous flaps and breast implants.

Some women electing immediate bilateral postmastectomy breast reconstruction have inadequate autologous tissue to achieve an aesthetic result and also wish to avoid tissue expansion or a back scar. These patients are candidates for reconstruction using free transverse rectus abdominis musculocutaneous (TRAM) flaps and breast implants. Since November 1993, 5 women have been reconstructed bilaterally with a combination of free TRAM flaps and adjustable saline breast implants. Each patient was young (age, 29-47 years) and of slender body habitus. During the procedure, the implants were placed in a submuscular pocket formed by the pectoralis major, the serratus anterior, and the muscular portion of the TRAM flap. Careful approximation of the serratus anterior to the lateral border of the pectoralis major muscle created a reliable layer of tissue to protect the vascular pedicle of the TRAM flap from contact with the implant. The saline chambers of the implants were filled to 60% to 80% capacity at the time of surgery. TRAM flap skin was tailored to precisely replace missing breast envelope, and subcutaneous fat increased implant coverage, creating a mound with normal-appearing ptosis. There were no occurrences of microvascular thrombosis. Three patients experienced prolonged formation of serous fluid, with 2 patients requiring late aspiration of a serosa after drain removal. Although there were no documented infections, 1 patient experienced postoperative erythema of the mastectomy skin flaps and was treated empirically with antibiotics with resolution of the erythema. All breasts have remained soft and appear natural. Using free TRAM flaps and adjustable saline breast implants allows selected women to undergo immediate breast reconstruction without the need for prolonged tissue expansion or back scars. Aesthetic results are excellent and the immediate use of an implant does not appear to pose a risk to the success of the free tissue transfer.

Adult↗

PubMed1996

Comparison of resource costs between implant-based and TRAM flap breast reconstruction.

Resource costs, as measured by hours of time in the operating room, days of stay in the hospital, and other costs of care, were evaluated for 240 patients who underwent mastectomy with immediate breast reconstruction using either TRAM flaps or breast implants at The University of Texas M. D. Anderson Cancer Center. To make costs comparable, only patients who completed reconstruction of the nipple were included. As expected, the initial resource costs of implant-based reconstruction were much lower than those of TRAM flap reconstruction. After correcting for patients whose reconstructions were unsuccessful and including the costs of surgery subsequent to the initial reconstruction, however, the cost advantage of implant-based reconstruction disappeared. If current trends continue, it is likely that with increased follow-up, the long-term resource costs of implant-based reconstructions will continue to increase, while those of autogenous tissue reconstructions will not. Autogenous breast reconstruction with the TRAM flap therefore appears to be more cost-effective, in terms of time as well as dollars, in the long run than reconstruction based on prosthetic implants.

Breast Implants↗

PubMed1996

Silicon tissue assays: a comparison of nonaugmented cadaveric and augmented patient levels.

Tissue silicon assays were performed on 10 nonaugmented cadavers and 25 augmented women to confirm our previous cadaveric data and to establish silicon levels at local and distant sites in augmented women undergoing explant and further reconstruction. All assays were performed by inductively coupled plasma atomic emissions spectroscopy (ICP-AES). Cadaveric tissues were sampled from six sites: liver, spleen, breast, nipple, axilla (soft tissue and nodes), and subcutaneous tissue (abdominal). Augmented women undergoing explant surgery had a portion of their implant capsule, breast tissue, and for those undergoing autogenous reconstruction, a portion of the autogenous donor site or distant tissue sampled. Twenty-four women had silicone implants; one had saline. Results revealed baseline silicon levels in all 10 non-augmented cadavers. These results were equivalent to our previous data, confirming the validity of these baseline data. Silicon tissue levels in the 25 augmented women revealed elevated levels within the implant capsule and surrounding breast tissue. However, silicon levels obtained from distant sites in augmented women were equivalent to the baseline nonaugmented cadaveric levels. There was no correlation between intact or ruptured implants and symptoms of collagen-vascular disease. In conclusion, study validates our original data of baseline silicon levels in nonaugmented cadavers. Contrary to what some may perceive as silicone floating throughout the body, we have found elevated levels around the implant only, and levels at distant tissue sites were equivalent to the baseline cadaveric data.

Abdomen↗

PubMed1996

Comparison of resource costs of free and conventional TRAM flap breast reconstruction.

Resource costs, which are the costs to the hospital of providing a service, were measured for 154 patients who underwent mastectomy and immediate breast reconstruction with TRAM flaps. Unilateral and bilateral reconstructions were evaluated separately. The resource costs required to perform mastectomy and reconstruction with free TRAM flaps were then compared with those required when conventional TRAM flaps were used. The mean total resource cost in the free TRAM group was slightly higher than in the conventional TRAM group, but the difference was small (4.1 percent) and not statistically significant (p = 0.290). The mean resource cost of performing bilateral mastectomy and reconstruction was higher than that of unilateral mastectomy and reconstruction, but the difference was only 5.0 percent (p = 0.046). This study shows that the cost to an institution of providing breast reconstruction with free TRAM flaps is not significantly higher than that of performing reconstruction with conventional TRAM flaps. Also, our findings show that the resource costs of performing bilateral mastectomy and reconstruction are not much higher than those of treating only one breast.

Female↗

PubMed1996

Choice of flap and incidence of free flap success.

A review of 854 consecutive free flaps was performed to determine whether the choice of flap used for the reconstruction influenced the probability of a successful outcome. Flaps were grouped into nine categories: rectus abdominis, free transverse rectus abdominis myocutaneous, radial forearm, jejunum, latissimus dorsi, fibula, scapula, iliac crest, and other. There were significant differences among the success rates of different flaps (p < 0.0001). Rectus abdominis-based flaps used for breast or head and neck reconstruction had lower failure rates (0.9 percent) than did non-rectus abdominis flaps (6.6 percent; p < 0.0001). Flaps requiring vein grafts had a higher rate of flap loss (18.4 percent) than did flaps that did not require vein grafts (2.9 percent; p < 0.0001). There was a strong trend favoring survival of flaps without a bone component (compared with osteocutaneous flaps), and a weaker trend favoring survival of flaps in nonobese patients (compared with flaps in obese patients). Smoking, age, and previous irradiation had no significant effect on flap failure rates. Surgeons should consider the flap success rate as one (but not necessarily the most important) factor in choosing the best reconstruction for any individual patient.

Breast Neoplasms↗

PubMed1996

Timing of pedicle thrombosis and flap loss after free-tissue transfer.

A series of 990 consecutive free flaps was reviewed to determine how often pedicle thrombosis occurred, when it occurred, and if the timing of thrombosis detection had any relationship to the probability of flap salvage. The overall thrombosis rate was 5.1 percent, and the flap loss rate was 3.2 percent. The majority (80 percent) of thrombi occurred within the first 2 postoperative days. Only 5 thrombi (10 percent) were known to have occurred after the third postoperative day. No flaps that developed thrombosis after the third postoperative day were salvaged successfully. Had flap monitoring been discontinued after the first 3 postoperative days, our results in this series would have been unchanged. Thrombosis of the vein (54 percent) was more common than arterial thrombosis (20 percent) or thrombosis of both artery and vein (12 percent). Almost all purely arterial thrombi (90 percent) occurred before the end of the first postoperative day, whereas 41 percent of all venous thrombi occurred later. We conclude that arterial monitoring is most critical immediately after surgery. Beginning on the second postoperative day, venous monitoring becomes progressively more important to flap success. The cost-effectiveness of postoperative monitoring of free flaps is greatest during the first 2 days, after which it decreases significantly.

Graft Rejection↗

PubMed1996

Species selectivity of new siderophore-drug conjugates that use specific iron uptake for entry into bacteria.

Siderophores selectively bind ferric iron and are involved in receptor-specific iron transport into bacteria. Several types of siderophores were synthesized, and growth-promoting or inhibitory activities when they were conjugated to carbacephalosporin, erythromycylamine, or nalidixic acid were investigated. Overall, 11 types of siderophores and 21 drug conjugates were tested against seven different bacterial species: Escherichia coli, Bordetella bronchiseptica, Pasteurella multocida, Pasteurella haemolytica, Streptococcus suis, Staphylococcus aureus, and Staphylococcus epidermidis. In some species, the inhibitory activities of the drug conjugates were associated with the ability of the bacteria to use the siderophore portion of the molecules for growth promotion in disc diffusion tests (0.04 mumol of conjugate or siderophore per disc). E. coli used catechol-based siderophore portions as well as hydroxamate-based tri-delta-OH-N-OH-delta-N-acetyl-L-ornithine ferric iron ligands for growth under iron-restricted conditions achieved by supplemental ethylenediamine di (O-hydroxyphenylacetic acid) (100 micrograms/ml) and was sensitive to carbacephalosporin conjugated to these siderophore types (up to a 34-mm-diameter inhibition zone). B. bronchiseptica used desferrioxamine B and an isocyanurate-based or trihydroxamate in addition to catechol-based siderophore portions for promotion but was not inhibited by beta-lactam conjugates partly because of the presence of beta-lactamase. P. multocida and P. haemolytica did not use any of the synthetic siderophores for growth promotion, and the inhibitory activities of some conjugates seemed partly linked to their ability to withhold iron from these bacteria, since individual siderophore portions showed some antibacterial effects. Individual siderophores did not promote S. suis growth in restrictive conditions, but the type of ferric iron ligands attached to beta-lactams affected inhibitory activities. The antibacterial activities of the intracellular-acting agents erythromycylamine and nalidixic acid were reduced or lost, even against S. aureus and S. epidermidis, when the agents were conjugated to siderophores. Conjugate-resistant E. coli mutants showed the absence of some iron-regulated outer membrane proteins in gel electrophoresis profiles and in specific phage or colicin sensitivity tests, implying that the drugs used outer membrane receptors of ferric complexes to get into cells.

Animals↗

PubMed1996

Growth of Actinobacillus pleuropneumoniae is promoted by exogenous hydroxamate and catechol siderophores.

Siderophores bind ferric ions and are involved in receptor-specific iron transport into bacteria. Six types of siderophores were tested against strains representing the 12 different serotypes of Actinobacillus pleuropneumoniae. Ferrichrome and bis-catechol-based siderophores showed strong growth-promoting activities for A. pleuropneumoniae in a disk diffusion assay. Most strains of A. pleuropneumoniae tested were able to use ferrichrome (21 of 22 or 95%), ferrichrome A (20 of 22 or 90%), and lysine-based bis-catechol (20 of 22 or 90%), while growth of 36% (8 of 22) was promoted by a synthetic hydroxamate, N5-acetyl-N5-hydroxy-L-ornithine tripeptide. A. pleuropneumoniae serotype 1 (strain FMV 87-682) and serotype 5 (strain 2245) exhibited a distinct yellow halo around colonies on Chrome Azurol S agar plates, suggesting that both strains can produce an iron chelator (siderophore) in response to iron stress. The siderophore was found to be neither a phenolate nor a hydroxamate by the chemical tests of Arnow and Csaky, respectively. This is the first report demonstrating the production of an iron chelator and the use of exogenous siderophores by A. pleuropneumoniae. A spermidine-based bis-catechol siderophore conjugated to a carbacephalosporin was shown to inhibit growth of A. pleuropneumoniae. A siderophore-antibiotic-resistant strain was isolated and shown to have lost the ability to use ferrichrome, synthetic hydroxamate, or catechol-based siderophores when grown under conditions of iron restriction. This observation indicated that a common iron uptake pathway, or a common intermediate, for hydroxamate- and catechol-based siderophores may exist in A. pleuropneumoniae.

Actinobacillus pleuropneumoniae↗