The value of biomedical research training for veterinary anatomic and clinical pathologists.

Veterinary pathologists traditionally have been actively engaged in research as principal investigators and as collaborators. Pathologists frequently obtain advanced training in research; however, it appears that in the last 10 years there has been a reversal of a previous trend toward increasing numbers of pathologists obtaining PhD degrees. This has arisen despite an established shortage of veterinarians engaged in research. This article evaluates the benefits of research training for individual pathologists, including a wide spectrum of professional opportunities and additional skill development beyond that usually provided by diagnostic pathology training alone. Various training models are discussed, including combined and sequential diagnostic residency and research degree training as well as the nondegree research fellowship programs more commonly pursued in human medicine. Best-practice recommendations for program infrastructure, mentorship, time management, and a team approach to research and research training are advocated to facilitate the development of successful programs and to encourage a continued emphasis on integrated training for pathologists as both clinical diagnosticians and experimentalists. This article is intended to help prospective and active pathology trainees, their mentors, and educational administrators optimize opportunities to ensure the future vitality of veterinary pathologists, and their contributions, in basic and applied research.

Quantitative analysis of flavonol glycosides in Ginkgo biloba: a comparison of two analytical methods.

A comparison of testing methods for constituents of Ginkgo biloba was performed as part of the process for determining which of the many available analytical methods was most appropriate for inclusion in the American Herbal Pharmacopoeia (AHP). Three primary methods were reviewed: those adopted by the European Pharmacopoeia, the National Science Foundation-Institute for Nutraceutical Advancement (NSF-INA) and the United States Pharmacopeia. Methods were compared by evaluating sample preparation and hydrolysis, precision and methods of total flavonol glycoside calculation by two independent laboratories. The AHP adopted the method of NSF-INA for inclusion in the AHP monograph owing to observed superiority of this method in terms of precision, glycoside calculation, ease of sample preparation and chromatographic parameters.

Expression of cyclooxygenase-2 in the macula densa of human kidney in hypertension, congestive heart failure, and diabetic nephropathy.

Cyclooxygenase-2 (COX-2) is constitutively expressed in the macula densa of several laboratory animal species where it is considered to play a physiologic role in the regulation of basal renal function. Pertubations to normal homeostasis is shown to be associated with the upregulation of COX-2 in the macula densa of rats and dogs. In contrast, COX-2 has not been detected in the macula densa of normal adult human and non-human primate kidneys, suggesting a less prominent role of this isoform in normal renal function in these species. In this study, we characterized COX-2 expression in human kidneys collected from subjects with a clinical history indicative of compromised renal function associated with diabetic nephropathy (DN), hypertension, and congestive heart failure (CHF). COX-2 expression was evaluated by immunohistochemistry using isoform-specific antibodies and in situ hybridization. No COX-2 protein or mRNA was observed in the macula densa of normal kidneys (n= 11), whereas slight to moderate COX-2 expression was present in the macula densa of 7/15 subjects (46%) with DN, 5/11 (46%) subjects with hypertension, and 3/10 subjects (30%) with CHF. These results indicate that COX-2 is variably induced in the macula densa of the human kidney in compromised renal conditions and that COX-2-mediated prostaglandins may be involved in maintaining adequate renal functions in some patients with DN, hypertension, and CHF. This variability may be related to individual clinical status or synthesis of vasodilatory prostaglandins by cyclooxygenase-1 (COX-1).

Silicon-29 diffusion-ordered NMR spectroscopy (DOSY) as a tool for studying aqueous silicates.

The first use of silicon-29 diffusion-ordered NMR spectroscopy (DOSY) is reported, in a study of the speciation of aqueous silicates.

A space group assignment of ZrP2O7 obtained by 31P solid state NMR.

2-D 31P dipolar recoupling magic angle spinning NMR has been used to determine the true symmetry of the low temperature structure of ZrP2O7 for the first time.

Cyclooxygenase-2 expression in inflammatory lung lesions of nonhuman primates.

Mammalian cells contain two related but unique isoforms of cyclooxygenase (COX-1 and COX-2). COX-1 is expressed constitutively in a majority of tissues and is involved in the production of prostaglandins (PGs) that modulate normal physiologic functions. COX-2 is inducible by various stimuli and is involved in the production of PGs that modulate physiologic events in development, cell growth, and inflammation. With the exception of peribronchial glands and chondrocytes of peribronchial cartilage, COX-2 is not detectable in the normal lung of nonhuman primates. We evaluated COX-2 expression by immunohistochemical methods in the inflammatory lesions of two cynomolgus monkeys (Macaca fascicularis) with acute severe pneumonia. Both monkeys exhibited acute severe bronchopneumonia; histologically, lung lesions were characterized by infiltration of large numbers of neutrophils and fewer macrophages, mild bronchial epithelial hyperplasia, and slight type-2 pneumocyte hyperplasia. In both monkeys, mild to marked COX-2 immunoreactivity was detected within the cytoplasm of macrophages, bronchial epithelial cells, type-2 pneumocytes, and endothelial cells of blood vessels. No COX-2 immunoreactivity was detectable in the neutrophils that constituted >90% of the inflammatory cells. These observations suggest that in acute inflammatory lung lesions in nonhuman primates 1) COX-2 is induced in the bronchial and alveolar epithelial cells, 2) macrophages are the primary inflammatory cells that exhibit COX-2, and 3) neutrophils do not express COX-2.

Expression of cyclooxygenase-2 in transitional cell carcinoma of the urinary bladder in dogs.

OBJECTIVE: To evaluate expression of cyclooxygenase (COX)-1 and COX-2 in the urinary bladder epithelium of clinically normal dogs and in transitional cell carcinoma cells of dogs. ANIMALS: 21 dogs with transitional cell carcinoma of the urinary bladder and 8 dogs with clinically normal urinary bladders. PROCEDURE: COX-1 and COX-2 were evaluated by use of isoform-specific antibodies with standard immunohistochemical methods. RESULT: COX-1, but not COX-2, was constitutively expressed in normal urinary bladder epithelium; however, COX-2 was expressed in neoplastic epithelium in primary tumors and in metastatic lesions of all 21 dogs and in new proliferating blood vessels in 3 dogs. Also, COX-1 was expressed in the neoplastic cells. CONCLUSIONS AND CLINICAL RELEVANCE: Lack of expression of COX-2 in normal bladder epithelium and its substantial expression in transitional cell carcinoma cells suggest that this isoform may be involved in tumor cell growth. Inhibition of COX-2 is a likely mechanism of the antineoplastic effects of non steroidal antiinflammatory drugs.

Carbon-13 and fluorine-19 NMR spectroscopy of the supramolecular solid p-tert-butylcalix(4)arene.alpha,alpha,alpha-trifluorotoluene.

The supramolecular 1:1 host-guest inclusion compound, p-tert-butylcalix[4]arene x alpha,alpha,alpha-trifluorotoluene, 1, is characterized by 19F and 13C solid-state NMR spectroscopy. Whereas the 13C NMR spectra are easily interpreted in the context of earlier work on similar host-guest compounds, the 15F NMR spectra of solid 1 are, initially, more difficult to understand. The 19F[1H] NMR spectrum obtained under cross-polarization and magic-angle spinning conditions shows a single isotropic resonance with a significant spinning sideband manifold. The static 19F[1H] CP NMR spectrum consists of a powder pattern dominated by the contributions of the anisotropic chemical shift and the homonuclear dipolar interactions. The 19F MREV-8 experiment, which minimizes the 19F-19F dipolar contribution, helps to identify the chemical shift contribution as an axial lineshape. The full static 19F[1H] CP NMR spectrum is analysed using subspectral analysis and subsequently simulated as a function of the 19F-19F internuclear distance (D(FF) = 2.25 +/- 0.01 A) of the rapidly rotating CF3 group without including contributions from additional libration motions and the anisotropy in the scalar tensor. The shielding span is found to be 56 ppm. The width of the centerband in the 19F[1H] sample-spinning CP NMR spectrum is very sensitive to the angle between the rotor and the magnetic field. Compound 1 is thus an attractive standard for setting the magic angle for NMR probes containing a fluorine channel with a proton-decoupling facility.

Sulfathiazole polymorphism studied by magic-angle spinning NMR.

The literature on sulfathiazole polymorphs has many confusions and inconsistencies. These are largely resolved by the distinctive appearance of (13)C magic-angle spinning NMR spectra, which immediately show the number of molecules in the crystallographic asymmetric unit. The spectra presented include those of a newly-recognized form. The assignments of the spectra are established and discussed in relation to such factors as electronic structure of the aromatic ring, second-order quadrupolar effects originating from the nitrogen nuclei, and hydrogen bonding. The results are compared to literature information on the crystal structures. When the amino group acts as a hydrogen bond acceptor, there is a shielding effect on C-4 to the extent of ca. 8 ppm (which should be compared to a further shielding by ca. 10 ppm for sulfathiazole sulfate). The fact that the spectrum of form III is similar to the sum of those of forms IV and V is rationalized in relation to the crystal structures. Some surprising variability of spectra with temperature and with specific sample is reported.

6Li and 7li solid-state NMR spectroscopy of nitrogen ceramic phases.

Two nitrogen ceramic phases, the oxynitride LiSiON and the nitride LiSi2N3, have been studied by 6Li and 7Li NMR. Magic angle spinning (MAS) NMR experiments have been carried out at two magnetic field strengths (7.05 and 14.1 T). The spectra give evidence of the relative effects of the quadrupolar and chemical shift interactions. The electric field gradient tensor of both phases has been determined accurately by iterative fitting of the 6Li and 7Li MAS NMR line shapes at the two magnetic field strengths. Due to the fact that for 7Li the quadrupolar interaction is much larger than the chemical shift interaction, it is shown that neither the small chemical shift anisotropy nor the relative orientation of the two interaction tensors can be determined accurately by 7Li MAS NMR. For 6Li, the two interactions are comparable and the value of these parameters obtained from the fits of the 6Li experimental MAS line shapes are therefore much more reliable.

A method for analysing proton NMR relaxation data from motionally heterogenous polymer systems.

Historically, the results of studies of the motional processes present in polymers above the glass transition temperature (Tg) by proton NMR spin-lattice relaxation in either the laboratory or rotating frames (T1 or T1p) have shown poor agreement with the results from similar studies carried out using other techniques such as dielectric or mechanical relaxation. We believe that this is mainly because of the complication of the NMR results due to magnetisation transport, either by spin diffusion or by bulk diffusion of the polymer. We suggest a novel approach to the analysis of proton NMR relaxation data from a motionally heterogeneous polymer, and show that the results of such an analysis are intrinsically reasonable and of the form expected for dielectric or mechanical relaxation.

Parameters and symbols for use in nuclear magnetic resonance (IUPAC recommendations 1997).

NMR is now frequently the technique of choice for the determination of chemical structure in solution. Its uses also span structure in solids and mobility at the molecular level in all phases. The research literature in the subject is vast and ever-increasing. Unfortunately, many articles do not contain sufficient information for experiments to be repeated elsewhere, and there are many variations in the usage of symbols for the same physical quantity. It is the aim of the present recommendations to provide simple check-lists that will enable such problems to be minimised in a way that is consistent with general IUPAC formulation. The area of medical NMR and imaging is not specifically addressed in these recommendations, which are principally aimed at the mainstream use of NMR by chemists (of all sub-disciplines) and by many physicists, biologists, materials scientists and geologists etc. working with NMR. The document presents recommended notation for use in journal publications involving a significant contribution of nuclear magnetic resonance (NMR) spectroscopy. The recommendations are in two parts: (1) Experimental parameters which should be listed so that the work in question can be repeated elsewhere. (2) A list of symbols (using Roman or Greek characters) to be used for quantities relevant to NMR.

Solid-state 19F NMR investigation of poly(vinylidene fluoride) with high-power proton decoupling.

The use of high-power proton decoupling has enabled highly-resolved spectra of fluorine polymers to be recorded, as is exemplified herein for semicrystalline poly(vinylidene fluoride) (PVDF). By means of high MAS speeds (up to 17 kHz), the spinning sidebands are removed from the whole of the relevant chemical shift range. For spectra of the crystalline regions of the polymer, the high-power decoupling is necessary, though its effect is not large. Various relaxation techniques have been used to examine the semicrystallinity and the polymorphism of PVDF, with special pulse sequences used to discriminate between the various domains. Different chemical shifts have been observed for the signals of the amorphous and crystalline phases. Those of the more immobile parts cover a substantial range.

Effect of 1H-decoupling in two-dimensional multiple-quantum MAS NMR spectroscopy of 23Na in a hydrous layered silicate.

In order to check the efficiency of high-power 1H decoupling when used with the two-dimensional multiple-quantum MAS method recently proposed to obtain isotropic spectra from quadrupolar nuclei, we applied this new technique to 23Na NMR for a hydrous layered silicate compound, makatite, with and without 1H decoupling. A remarkable improvement of the spectral resolution in the isotropic dimension was observed by the decoupling, showing that it is effective in the 2D MQ-MAS method and suggesting that proton decoupling should be generally applied in 23Na experiments concerning these kinds of compound.

Respiratory and pharyngo-esophageal iridovirus infection in a gopher tortoise (Gopherus polyphemus).

A free-living adult male gopher tortoise (Gopherus polyphemus) was found on Sanibel Island, Florida (USA), on 18 February 1992 with signs of upper respiratory disease. On necropsy after euthanasia on 27 February 1992, severe, extensive necrotizing ulcerative tracheitis, multifocal necrotizing pneumonia, and multifocal necrotizing ulcerative pharyngitis and esophagitis were observed. Large ovoid to round intracytoplasmic basophilic inclusions, which appeared to displace the nucleus to the cell periphery, occurred within degenerate and necrotic epithelial cells of the above tissues. On transmission electron microscopy of formalin-fixed trachea and lung, intracytoplasmic viral particles were observed within necrotic cells in the tracheal lumen and epithelial cells of the lung. Most infected cells also had a roughly spherical granular cytoplasmic inclusion that contained clusters of viral particles. Viral particles had an electron dense spherical to icosahedral core surrounded by a less electron dense icosahedral capsid. Mature extracellular virions were surrounded by an envelope and were 150 to 220 nm in diameter. Virions and cytoplasmic inclusions were morphologically similar to those of the Family Iridoviridae.

Proton fluorine wideline separation experiments on poly (vinylidene difluoride).

Two-dimensional heteronuclear proton-fluorine correlation solid-state spectroscopy is demonstrated on a sample of poly (vinylidene difluoride) (PVDF). This allows the correlation of structural information contained in the fluorine chemical shift in one dimension with information on mobility from proton wideline spectra in the second dimension. The effect of the heteronuclear fluorine-proton dipolar coupling on the proton linewidth is investigated separately. The incorporation of magnetization filters into the preparation period enables one to measure the wideline separation (WISE) spectra for the amorphous or the crystalline parts of the sample separately.

1H, 29Si and 1H, 23Na heteronuclear shift correlation in octosilicate via cross-polarisation.

Solid-state magic-angle spinning was used to study octosilicate, a layered sodium polysilicate hydrate. Conventional one-dimensional NMR spectra detect two distinct proton and silicon sites and a single sodium species. Heteronuclear shift correlation experiments have shown that only one type of proton is involved in efficient cross-polarisation to all the silicon and sodium sites. Such selectivity could be caused by a favourable position for, and suitable rigidity of, this strongly hydrogen-bonded proton. The latter requirement can be used to propose a simple model of the interlayer space in octosilicate.

Clinical and clinical laboratory correlates in sea otters dying unexpectedly in rehabilitation centers following the Exxon Valdez oil spill.

Following the Exxon Valdez oil spill, 347 oiled sea otters (Enhydra lutris) were treated in rehabilitation centers. Of these, 116 died, 94 within 10 days of presentation. Clinical records of 21 otters dying during the first 10 days of rehabilitation were reviewed to define the laboratory abnormalities and clinical syndromes associated with these unexpected deaths. The most common terminal syndrome was shock characterized by hypothermia, lethargy, and often hemorrhagic diarrhea. In heavily and moderately oiled otters, shock developed within 48 hours of initial presentation, whereas in lightly oiled otters shock generally occurred during the second week of captivity. Accompanying laboratory abnormalities included leukopenia with increased numbers of immature neutrophils (degenerative left shift), lymphopenia, anemia, azotemia (primarily prerenal), hyperkalemia, hypoproteinemia/hypoalbuminemia, elevations of serum transaminases, and hypoglycemia. Shock associated with hemorrhagic diarrhea probably occurred either as a direct primary effect of oiling or as an indirect effect secondary to confinement and handling in the rehabilitation centers. Lightly oiled otters were less likely to die from shock than were heavily oiled otters (22% vs. 72%, respectively). Heavily oiled otters developed shock more rapidly and had greater numbers of laboratory abnormalities, suggesting that exposure to oil was an important contributing factor.