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Evaluation of the microcirculation of the equine jejunum and ascending colon after ischemia and reperfusion
1993
Dabareiner, R.M. | Snyder, J.R. | Sullins, K.E. | White, N.A. II. | Gardner, I.A.
Intramural vascular patterns of the jejunum and colon were evaluated during ischemic strangulation obstruction (ISO, 70 minutes) and subsequent reperfusion (60 minutes) in 7 adult anesthetized horses. Microvasculature of experimental and control segments was described by comparison of results from microangiography, light microscopy, and scanning electron microscopy of vascular replicas. Experimental and control segments with isolated vascular arcades were removed either immediately after the experimental period or after 60 minutes of reperfusion. Blood was flushed from the vascular system by use of isotonic NaCl, and the segments were divided. Half of each segment was perfused with a modified radiopaque medium for microangiographic evaluation, and half was perfused with dilute methylmethacrylate to create a vascular replica to be studied by scanning electron microscopy. Microangiographic section also were evaluated for histologic changes. Microvasculature of jejunal control segments and all colon segments was similar to described normal microvasculature of the equine jejunum and ascending colon. In jejunal ISO segments, intramural perfusion was redistributed away from the mucosa. In the villi, the central arteriole was short and convoluted and the subepithelial capillaries were not filled. The submucosal vessels and crypt capillaries were congested, compared with those of controls, and the serosal vessels were not filled in the ischemic segments. Histologic grade II-III mucosal lesion was seen in jejunal ISO segments. Reperfused jejunal segments had a transmural hyperemic response, and previously unfilled capillaries were observed in all intestinal layers. After reperfusion, the mucosal lesion progressed to grade III-IV and a cellular infiltrate and edema formation were observed in the serosa. The intramural vasculature of the ischemic and reperfused colon remain unchanged. Minimal histologic damage was observed in the colon after 70 minutes of ISO or after 60 minutes of reperfusion.
Показать больше [+] Меньше [-]Histomorphometric analysis of the omasum of sheep during development
1993
Franco, A. | Robina, A. | Regodon, S. | Vivo, J.M. | Masot, A.J. | Redondo, E.
Histomorphometric and scanning electron microscopic analyses were performed on 74 embryos and fetuses and 20 sheep (early postnatal to adult age). Histologic differentiation of the omasum took place at 33 days of fetal life, with the appearance of first-order laminae. Second-, third-, and fourth-order laminae appeared at 39, 50, and 59 days, respectively. Neutral mucopolysaccharides first appeared in epithelial cells at 46 days of fetal life, decreasing quantitatively until birth, before subsequently stabilizing in postnatal life. Acid mucopolysaccharides, mucins, and mucoid compounds were not detected. Growth curves and formulas were constructed for each tissue layer. Initial tests involved multiplicative (y = axb), exponential (y = EXP [a + bx]), linear (y = a + bx), and polynomial models [y = a + bx + cx(2) + dx(3)].
Показать больше [+] Меньше [-]Flow cytometric analysis of neutrophils in cows' milk
1993
Miller, R.H. | Paape, M.J. | Filep, R. | Link, S.
Procedures were developed to count neutrophils in milk using a flow cytometer. Milk samples from 2 experiments were counted: 1 with 4 noninfected cows and a second with 5 noninfected cows that were injected with endotoxin in 2 mammary quarters. Thus, the procedures were evaluated on normal milk and on that with high somatic cell count. Flow cytometric procedures involved fluorescence detection (from the dye carboxydimethylfluorescein diacetate) to distinguish intact and viable from fragmented cells, forward light scatter to detect cell size differences, and right-angle side scatter to detect cellular granularity. High fluorescence, large size, and high degree of granularity identified viable neutrophils. For all samples, neutrophils were also counted manually, using the cytologic centrifugation approach to create the slides; manual counts were used as the standard for comparison. In experiment 1 (normal milk), mean values for percentage of viable neutrophils estimated by manual and flow cytometry procedures agreed closely (26% vs 25.8% for foremilk and 28.8% vs 26.6% for bucket milk). Sources of variation in manual and flow cytometric estimates of percentage of neutrophils were examined. Cow variation was significant (P < 0.01) for manual and flow cytometric counts, but was larger for flow cytometric counts. Day-to-day variation in counts on milk from the same cow was negligible for manual counts, but was significant (P < 0.01) for flow cytometric counts. Coefficients of variation were considerably Larger for manual counts than for flow cytometry. In experiment 2 (milk with high cell count, foremilk), agreement between mean values obtained by flow cytometry and by manual counting was somewhat less. However, predicting manual percentage of neutrophils, using the flow cytometric estimate, had R2 of 0.77. Regression of the manual percentage of neutrophils value on the flow cytometric percentage of neutrophil value was close to 1.0, with only a small negative intercept. Some additional refinement of flow cytometric procedures may be required before flow cytometric estimates of percentage of neutrophils can be accepted without simultaneous validation by use of manual counting. In particular, causes of day-to-day variation in flow cytometric results should be identified and reduced.
Показать больше [+] Меньше [-]Ultrastructural hepatocellular features associated with severe hepatic lipidosis in cats
1993
Center, S.A. | Guida, L. | Zanelli, M.J. | Dougherty, E. | Cummings, J. | King, J.
In this study, we compared hepatic ultrastructure in healthy cats, in cats with severe hepatic lipidosis, and in cats with experimentally induced, chronic, extrahepatic bile duct occlusion. Ultrastructural features unique to the lipidosis syndrome included an apparent reduction in number of peroxisomes and alteration in their morphologic features. The quantity of endoplasmic reticulum, Golgi complexes, and lysosomes was subjectively reduced, and paucity of cytosolic glycogen was observed. Bile canaliculi appeared collapsed because of cytosolic distention with lipid. Mitochondria were reduced in number and were markedly pleomorphic. Cristae assumed a variety of shapes, lengths, and orientations. Ultrastructural features of bile duct occlusion were similar to those described in other species and differed from those in cats with hepatic lipidosis.
Показать больше [+] Меньше [-]Automated morphometric analysis of stallion spermatozoa
1993
Davis, R.O. | Gravance, C.G. | Casey, P.J.
Tissue variation in microscope slides made for spermatozoon analysis and variation introduced by the subjective techniques used to analyze these slides reduce the statistical power of studies that seek to use spermatozoon morphology to predict fertility. A simple specimen preparation method was developed to standardize stallion spermatozoon morphologic smears, and a new, automated spermatozoa morphometry instrument was used to objectively analyze the efficacy of the specimen preparation technique. The method achieved a standard spermatozoon concentration and reduced field-to-field variation in the number of spermatozoa analyzed. Metric measurements of spermatozoon head dimensions from clinically normal, fertile stallions revealed small, but highly significant, differences between stallions. The variation in metric measurements between replicate slides within stallions was small, indicating that replicate slide analysis probably is not necessary for clinically normal stallions. Coefficients of variation were generally less than 11% for metric measurements between stallions, and were less than 4% within stallions. This study revealed that a high degree of statistical power can be achieved when using these new, standardized specimen preparation and objective analysis techniques. Such power makes possible the detection of subtle differences between clinically normal stallions, and may facilitate accurate detection of abnormal fertility (ie, subfertility) in stallions.
Показать больше [+] Меньше [-]Ultrastructural features and pathogenesis of knobbed spermatozoa in a boar
1993
Toyama, Y. | Itoh, Y.
Ultrastructures of knobbed spermatozoa in a boar were observed. The knobs, found at the apex of the spermatozoa, were spherical swellings of the acrosome; vacuoles were found in the swellings. According to the contents, 2 types of the vacuoles were recognized: a vacuole containing cell debris that was surrounded by 2 or 3 layers of membranes, and a vacuole containing an amorphous material that was surrounded by a single membrane. Several vacuoles might be observed in a knob. Observations of both testes indicated that the cell debris in the vacuole of the knob was derived from the cytoplasm of the Sertoli cell, which evaginated into the spermatid. Origin of the amorphous material in the other type of knob is not known.
Показать больше [+] Меньше [-]Effect of 4-bromo-calcium ionophore A23187 on release of Anaplasma marginale from bovine erythrocytes in vitro
1993
The ionophore A23187 was used to facilitate release and continued development of Anaplasma marginale in short-term erythrocyte cultures. Addition of 10 micromolar A23187 to the cultures resulted in significant decrease in percentage of parasitized erythrocytes (PPE) by 24 hours after treatment; further development and increase in PPE was not observed. In contrast, the PPE of untreated cultures, those treated with dimethyl sulfoxide (DMSO) only and with 1 micromolar A23187 increased slightly during that time. Total erythrocyte count decreased in treated cultures in excess of that expected after samples of the medium were taken for analysis. The greatest cell loss and increased hemoglobin concentration in culture medium was observed in cultures treated with 10 micromolar A23187 and with an equivalent volume of DMSO. The DMSO appeared to cause hemolysis of some erythrocytes, but not of infected cells selectively. Release of A. marginale inclusion bodies was seen by electron microscopy in samples from the 10 micromolar A23187-exposed cultures. At 30 minutes after treatment, free initial bodies were frequently seen. Inclusion body membranes and individual A. marginale were associated with membranes of adjacent erythrocytes. Individual rickettsiae were seen in cell depressions and appeared to be entering erythrocytes. However, neither further invasion nor development of the parasite in erythrocytes was observed. Ionophore A23187 appeared to promote release of A. marginale from erythrocytes, but did not enhance infection of erythrocytes or development of organisms in vitro.
Показать больше [+] Меньше [-]Sequential study of pancreatic structure and function during development of pancreatic acinar atrophy in a German Shepherd Dog
1993
Westermarck, E. | Batt, R.M. | Vaillant, C. | Wiberg, M.
Sequential assessments of pancreatic structure and function were performed on a female German Shepherd Dog bred from parents with exocrine pancreatic insufficiency (EPI), to monitor development of pancreatic acinar atrophy in this breed. Determinations of serum trypsin-like immunoreactivity (TLI), results of N-benzoyl-L-tyrosyl-P-aminobenzoic acid test, fecal soy bean stimulation test (SST), and gross and histologic examinations of the pancreas did not provide evidence of exocrine pancreatic disease up to 13 months of age. However, electron microscopy revealed degenerative abnormalities of acinar cells that were already apparent at 6 weeks and became more extensive with age. Examination of the pancreas at 22 months of age also indicated no gross or histologic abnormalities, but electron microscopy revealed widespread degenerative changes, including dilatation of the rough endoplasmic reticulum and extensive fusion of zymogen granules affecting most of the acinar cells. Serum TLI concentration nm markedly reduced at that time, indicative of EPI, but the dog remained healthy and results of the SST were normal. Within 1 month, the dog had developed clinical signs of EPI, and not only serum Tli concentration, but also results of the N-benzoyl-L-tyrosyl-P-aminobenzoic acid test and SST were compatible with severe loss of exocrine pancreatic tissue. This loss was confirmed by gross and histologic examination of the pancreas at 25 months, which revealed typical features of pancreatic acinar atrophy, including scattered and disorganized exocrine cells in the small remnants of pancreatic tissue. These findings indicate that in German Shepherd Dogs, pancreatic acinar atrophy may involve interference with normal intracellular processing of
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