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PubMed1999

Human megakaryocyte ploidy.

We reviewed the literature concerning the history of determination of the ploidy of human megakaryocytes and its relationship with diseases. The ploidy of rabbit megakaryocytes was analyzed by microspectrophotometry in 1964, and the analysis of the ploidy in human megakaryocytes was first performed in 1968. Presently, microphotometry and flow cytometry are the primary methods for the evaluation of the ploidy, but they have their merits and demerits. In the ploidy of human megakaryocytes, a peak has often been reported at 16N in healthy individuals, and the next peaks have been observed at 32N and 8N. The results of ploidy analyses have been reported by many investigators to be comparable between patients with idiopathic thrombocytopenic purpura and normal subjects, but various shifts of the peaks have also been documented. The ploidy is often reported to shift to a larger ploidy class in polycythemia vera and essential thrombocythemia, but it has invariably been reported to shift to a smaller class in chronic myelogenous leukemia. In reactive thrombocytosis, the ploidy pattern was reported to be the same as that in normal individuals by some investigators but to shift to a larger ploidy by others. These differences are considered to be due to heterogeneity of the subjects. In myelodysplastic syndrome, the ploidy shifts mostly to a smaller class, but it may show various patterns. We also reviewed the ploidy in other rare hematological disorders, the relationships of the ploidy with diabetes mellitus and atherosclerotic disorders, and its changes in the ontogeny. Details of the mechanism of polyploidization and its biological significance remain unknown, and further advances in the studies of these topics are anticipated.

Animals↗
PubMed2002

Intracellular photobleaching of 5,10,15,20-tetrakis(m-hydroxyphenyl) chlorin (Foscan) exhibits a complex dependence on oxygen level and fluence rate.

The understanding of photosensitizer photobleaching is important not only for mechanistic studies, but also for the development of monitoring techniques for clinical dosimetry in photodynamic therapy. In this study, we investigated the intracellular photobleaching of 5,10,15,20-tetrakis(m-hydroxyphenyl)chlorin (mTHPC, Foscan) in the murine macrophage cell line J774A.1, using quantitative fluorescence imaging microscopy, microspectrofluorometry and microspectrophotometry. Using 652 nm laser irradiation, it was found that mTHPC exhibits oxygen- and fluence rate-dependent intracellular photobleaching. The kinetics showed an inverse dose-rate behavior, i.e. a reduction of fluence rate resulted in more photobleaching at comparable fluences. The effect of deoxygenation was found to be more complex, with decreased bleaching at low fluence rates and increased bleaching at higher fluence rates. The intracellular formation of reactive oxygen species was measured using 2',7'-dichlorodihydrofluorescein diacetate. The results are analyzed in terms of competitive Type-I and Type-II mechanisms.

Animals↗
PubMed1975

RNA content in the heart muscle cells following adrenalectomy and additional overload in the rat.

RNA content in the heart muscle cells of the adrenalectomized rats and those with aortic constriction was measured by microspectrophotometry. In the adrenalectomized rats, the RNA content decreased about 32% below that of the intact control rats by 7 days after operation. On the other hand, it decreased more abruptly in the adrenalectomized rats with aortic constriction. These results suggested that in the adrenalectomized rats, hypoplasia of the heart was caused by reduction of RNA content and myocardial compensatory hypertrophy was never induced even by overloading to the heart. The fact that the reduction of RNA content preceded the decrease of the heart weight per 100 g body weight suggests that in the cases of sudden unexpected death by unknown cause, the measurement of RNA content of heart muscle cells may be a clue to judge whether the cause of death is concerned with the heart or not.

Adrenal Cortex Hormones↗
PubMed1980

Cytological examination of cerebrospinal fluid in eight patients with neuro-Behçet's disease.

In eight patients with Neuro-Behçet's disease cerebrospinal fluid (CSF) cells were studied in relation to the clinical symptoms, clinical course and treatment. The results obtained were as follows: (1) The mean value of CSF cell counts was 126 +/- 177/3 mm3 in 88 specimens from the patients, and 2.9 +/- 2.4/3 mm3 in 30 specimens from healthy control subjects, the former being significantly higher than the latter (p less than 0.01). (2) In the patients' CSF, the average percentages of mononuclear and polymorphonuclear cells were 67 +/- 23% and 32 +/- 23%, respectively. (3) In CSF cytogram from 37 specimens from the patients, the average percentage of small lymphocytes was 59 +/- 19%, which was significantly lower (p less than 0.01) than control (71 +/- 8%). The average percentage of activated lymphocytes, activated monocytes and neutrophils which were not observed in control were 0.5 +/- 1.2%. 1.9 +/- 2.2% and 17 +/- 16%, respectively. (4) The ratio of nucleic acids (RNA/DNA) measured by microspectrophotometry (Zeiss UMSP-1) in CSF lymphocytes stained with methylgreen and pyronin was 0.58 +/- 0.12 in 17 specimens from the patients and 0.55 +/- 0.07% in 10 specimens from the control; there was no significant difference between them. These results suggested that the pleocytosis in Neuro-Behçet's disease may represent perivascular inflammatory process in the central nervous system.

Adult↗
PubMed1985

Electrophysiological measurements of the spectral sensitivity of three types of cones in the carp retina.

The spectral sensitivities of red-, green-, and blue-sensitive cones were measured by intracellular recording in the carp retina. The responses from all cones were univariant, i.e., the waveform of a response to any wavelength and at any intensity could be superimposed on the response to other wavelengths if the intensity was properly selected. Red-sensitive cones showed a maximum sensitivity at about 620 nm, green-sensitive cones at about 520 nm, and blue-sensitive cones at about 460 nm. The peak wavelengths and general forms of the spectral sensitivity curves agreed with those of the spectral absorption curves measured by microspectrophotometry (MSP), but the sensitivity of red-sensitive cones at both ends of the spectrum was significantly lower than the spectral absorption determined by MSP.

Animals↗
PubMed2002

Intravascular inhomogeneous distribution of oxygen in arterioles of rat cremaster muscle.

Previously, cross-sectional oxygen distribution in microvessels was assumed to be homogeneous. The oxygen profile in the arterioles of rat cremaster muscle was measured using microspectrophotometry and a PO(2) microelectrode, showing a drop in SO(2) as well as PO(2) close to the vascular wall with a flat PO(2) profile in the perivascular tissue.

Abdominal Muscles↗
PubMed1999

Spectral sensitivity of vision and bioluminescence in the midwater shrimp Sergestes similis

In the oceanic midwater environment, many fish, squid, and shrimp use luminescent countershading to remain cryptic to silhouette-scanning predators. The mid-water penaeid shrimp, Sergestes similis Hansen, responds to downward-directed light with a dim bioluminescence that dynamically matches the spectral radiance of oceanic down-welling light at depth. Although the sensory basis of luminescent countershading behavior is visual, the relationship between visual and behavioral sensitivity is poorly understood. In this study, visual spectral sensitivity, based on microspectrophotometry and electrophysiological measurements of photoreceptor response, is directly compared to the behavioral spectral efficiency of luminescent countershading. Microspectrophotometric measurements on single photoreceptors revealed only a single visual pigment with peak absorbance at 495 nm in the blue-green region of the spectrum. The peak electrophysiological spectral sensitivity of dark-adapted eyes was centered at about 500 nm. The spectral efficiency of luminescent countershading showed a broad peak from 480 to 520 nm. Both electrophysiological and behavioral data closely matched the normalized spectral absorptance curve of a rhodopsin with lambda(max) = 495 nm, when rhabdom length and photopigment specific absorbance were considered. The close coupling between visual spectral sensitivity and the spectral efficiency of luminescent countershading attests to the importance of bioluminescence as a camouflage strategy in this species.

Journal Article↗
PubMed1991

Rough ER degranulation and swelling resolved as lysosomes in Na+/H+ antiporter mediated rounding of human lung fibroblasts.

Na+/H+ antiporter activation by incubation in bicarbonate buffered saline induces long flat anchored human lung fibroblasts in monolayer culture to retract their cytoplasmic processes and become eventually globular in form. Reduction of cell profile area and perimeter in antiporter mediated rounding (AMR) is quantitated by microspectrophotometry and the roundness form factor (PE) calculated from these parameters. Quantitatively, 2 stages of rounding are discernible. Stage I rounding is a simple retraction phenomenon where PE upshifts marginally. Stage II rounding sees the cell transformed into globular refractile form where PE greater than 0.5. Ultrastructural studies show appreciable swelling of rough ER in Stage I rounding. Stage II rounding is accompanied by enhanced swelling of rough ER and degranulation. A dynamic relationship between rough ER (RER) and smooth ER (SER) in different functional states of a cell is novel. Enlarged SER channels appear resolved in later stages as lysosomal bodies suggesting that degranulated ER membranes are lysosomal targets in autophagocytosis.

Bicarbonates↗
PubMed1986

A comparison of internal standards for plant cytophotometry.

Plant cell nuclei were compared with chicken erythrocyte nuclei for use as internal standards for microspectrophotometry. The amount of DNA per nucleus and the coefficient of variation for measurement of individual nuclei were determined for cells from dormant embryos of Pinus taeda and Pinus coulteri, from onion root tips and from chicken erythrocytes. The chicken erythrocytes had the least variability and thus were best suited for use as a standard. Onion root tips were least suitable, with a coefficient of variation 2 1/2 times that of erythrocytes. Although onion root tips have been used as an internal standard in other studies, their mitotic activity, in contrast with the nonreplication of DNA of mature erythrocytes, is reflected in a broad distribution of nuclei with values in the 2C-4C range. Coulter pine mature embryos were at the 3C level, whether dry or hydrated, while loblolly pine embryos were in the 2C state. This confirms previous reports. The coefficient of variability for the pine embryo cells was 1 1/2 times that of erythrocytes for nonhydrated seeds and twice the erythrocyte value for hydrated seeds. The larger 2C values for pine (26 pg for P. taeda and 17 pg for P. coulteri) are closer to values expected for many plant species than the 3 pg level of the chicken erythrocytes. Dormant P. taeda embryo cells (2C) are suggested as an alternative where the experimental material has large DNA values and/or chicken erythrocytes are difficult to procure. Large sample size is recommended for the plant materials if they are to be used as internal standards in Feulgen cytophotometry.

Allium↗
PubMed1994

Changes in perimicrovascular protein spatial distribution due to superfusate.

OBJECTIVE: To determine superfusate-induced changes in the distribution of plasma proteins in the perimicrovascular interstitial matrix. METHODS: Rats were anesthetized with sodium pentobarbital and a mesenteric loop was exteriorized. Intravital video microspectrophotometry was performed using wavelengths of 280, 320, and 700 nm. The images were analyzed to give protein and collagen spatial distributions in vascular regions of rat mesenteric tissue. Perimicrovascular protein concentrations were fitted to an exponential decay model ci + cr exp (-x/k), where ci is distal protein concentration, ci + cr is the protein concentration proximal to the vessel, x is the distance from the vessel wall, and k is the decay constant indicating protein gradient slope. RESULTS: Before superfusion with 0.5-ml normal saline, ci = 1.45 +/- 0.13 g/dl, ci + cr = 4.56 +/- 0.23 g/dl. After the first superfusion, ci decreased (p < 0.01) to 0.53 +/- 0.06 g/dl; following a second superfusion, cr decreased to 0.4 +/- 0.03 g/dl; an additional final superfusion caused a further decrease to 0.33 +/- 0.02 g/dl. ci + cr also decreased significantly during repeated superfusions to 2.92 +/- 0.15, 2.35 +/- 0.25, and 2.1 +/- 0.12 g/dl, respectively. CONCLUSIONS: Superfusion produced changes in perivascular and distal interstitial matrix protein distribution. Protein concentration proximal to the microvessel remained higher than distal concentrations. This could be due to increased gel concentrations inhibiting protein mobility.

Animals↗
PubMedDate not supplied

Regional myocardial efficiency is improved in experimental aorto-caval shunt.

We determined whether regional myocardial work efficiency (segment work/regional O2 consumption) would be elevated by surgically-augmented inflow. In 10 anesthetized open-chest dogs, shunt between the ascending aorta and the superior vena cava was used to increase cardiac output. Hetastarch (15 ml/kg) was infused before opening the shunt to maintain coronary perfusion pressure. Regional myocardial segment work and O2 consumption (MVO2) were measured, during control and two levels of elevated flow. Regional segment work (g.mm/min) was calculated as the integrated products of force (g - miniature transducer) and segment shortening (mm - ultrasonic dimension gauge) during an averaged beat expressed per minute. Local MVO2 (ml O2/min/100g) was calculated from regional blood flow (microspheres) and O2 saturations (microspectrophotometry). It was found that regional myocardial segment work increased significantly (P < 0.05) from 926 +/- 94 to 1656 +/- 220 to 1479 +/- 309 (g.mm/min) for closed, half-open, and open shunt. This increase was primarily associated with increased segment shortening (from 147 +/- 14.1 to 204.4 +/- 20.1 to 232 +/- 26.1 mm/min). Both force development and regional MVO2 were unchanged during the experiment. Regional myocardial efficiency was significantly elevated during shunt function (from 95 +/- 12 to 187 +/- 31 to 213 +/- 57 g.mm/ml O2/100g). Systolic ejection stiffness (defined as the slope of the force-length relationship during the period of ejection) decreased from 8.0 +/- 0.9 to 4.7 +/- 0.4 to 4.5 +/- 0.9 g/mm during elevated inflow. It is concluded that when cardiac work is augmented primarily by segment shortening, regional myocardial efficiency is improved. This improvement is associated with decreased resistance to shortening (stiffness).

Animals↗
PubMed1988

DNA cytometry of osteosarcoma.

The relationship between cytochemical features and histomorphology in osteosarcoma, and the clinical significance of DNA content were investigated by microspectrophotometry (MSP) of tissue sections and flow cytophotometry (FCM) of cell suspensions. MSP of tissue sections entails the methodological error of determining the DNA content of sectioned cell nuclei. By analyzing 184 normal mesenchymal cell populations, an upper limit of diploidy (normal DNA content) was deduced. Applying this upper limit for 42 sarcomas, 6 were diploid and 36 hyperploid. Comparative analysis of the same lesions by MSP of imprint preparations and by FCM disclosed complete agreement in ploidy classification (diploid versus hyperploid). Retrospective MSP analysis of bone tumors is often impeded by previous demineralization in acid, which destroys DNA. EDTA as an alternative was found to slightly reduce Feulgen DNA stainability of osteosarcomas, but did not affect tumor ploidy determination. Hence, EDTA offers a means of retaining DNA stainability of bone tumors requiring demineralization. MSP analysis of different histologic areas, and comparative FCM analysis of biopsy and surgical specimens, dislosed that individual osteosarcomas are cytochemically uniform despite morphologic heterogeneity. Hence, a single tumor sample for DNA analysis can be relied upon as representative for the tumor as a whole. In a consecutive series of 83 osteosarcoma patients treated by surgery and adjuvant Interferon, the 7-year survival rate was 0.44. MSP DNA analysis gave no significant prognostic information. Multivariate analysis identified 3 risk factors for tumor related death, i.e., male sex, proximal tumor location, and histologic grade IV. In a prognostication model, the 7-year survival rates, for patients with 0, 1, 2, or 3 risk factors, were 0.80, 0.59, 0.42, and 0.13, respectively. Hence, it is possible to identify subgroups of high grade osteosarcoma patients with different prognosis. In a study of 166 primary bone tumors, the applicability of DNA analysis for differential diagnostic purposes was investigated. The series included high grade osteosarcomas, parosteal osteosarcomas and benign bone tumors, which may be mixed up histologically with osteosarcoma. Out of 166 tumors, 149 (90%) were histologically noncontroversial, whereas 17 (10%) posed diagnostic difficulties. In the diagnostically noncontroversial group, all benign tumors and parosteal osteosarcomas were diploid, whereas 97 of 102 osteosarcomas were hyperploid. Hence, hyperploidy seems to be a characteristic feature of high grade osteosarcoma.(ABSTRACT TRUNCATED AT 400 WORDS)

Bone Neoplasms↗
PubMed2005

Effect of cholecystokinin on experimental neuronal aging.

AIM: To observe the effect of cholecystokinin (CCK) on lipofusin value, neuronal dendrite and spine ultrastructure, and total cellular protein during the process of experimental neuronal aging. METHODS: Experimental neuronal aging study model was established by NBA2 cellular serum-free culture method. By using single intracellular lipofusin value from microspectrophotometry, morphology of neuronal dendrites and spines from the scanner electron microscopy, and total cellular protein as the indexes of experimental neuronal aging, we observed the effect of CCK8 on the process of experimental neuronal aging. RESULTS: Under the condition of serum-free culture, intracellular fluorescence value (%) increased with the extension of culture time (1 d 8.51+/-3.43; 5 d 10.12+/-3.03; 10 d 20.54+/-10.3; 15 d 36.88+/-10.49; (b)P<0.01). When CCK was added to serum-free culture medium, intracellular lipofusin value (%) decreased remarkably after consecutive CCK reaction for 10 and 15 d (control 36.88+/-10.49; 5 d 32.03+/-10.01; 10 d 14.37+/-5.55; 15 d 17.31+/-4.80; (b)P<0.01). As the time of serum-free culturing was prolonged, the number of neuronal dendrite and spine cells decreased. The later increased in number when CCK8 was added. CCK8 could improve the total cellular protein in the process of experimental neuronal aging. CONCLUSION: CCK8 may prolong the process of experimental neuronal aging by maintaining the structure and the number of neuronal dendrite and spine cells and changing the total cellular protein.

Animals↗
PubMedDate not supplied

Glycoconjugates, cellular differentiation, and congenital glaucoma.

Studies of congenital glaucoma have described and debated the disease process in cellular and structural terms, yet the functional aspects of the disease remain poorly understood. This paper presents an attempt to look at the disease from a different perspective: the role of glycoconjugates and their influence on morphogenesis of the corneoscleral angle. Extracellular and cell-surface complex carbohydrates and trabecular meshwork development were observed in normal C-57 BL/6J mice with light and electron microscopy. Lectin-stained fixed tissues were evaluated by fluorescein light microscopy and with computer-controlled scanning microspectrophotometry. The appearance and type of specific glycoconjugates was related to morphogenesis and differentiation, with specific reference to the development of the trabecular meshwork. Our results suggest that glycoconjugates are: (1) important in the morphogenesis of the corneoscleral angle, and (2) provide some of the required signals for the differentiation of the cells of the trabecular meshwork. These findings emphasize the possibility that a complex series of morphological and biological events in the normal development of the trabecular meshwork ultimately leads to a functionally competent trabecular meshwork, that is, capacitation.

Animals↗
PubMed1976

[Microspectrophotometric study of oxidative enzyme activity in experimental myocardial ischemia].

The method of comparitive microspectrophotometry was used to study the changes in the activity of oxidative enzymes in the zone of ischaemia and the juxtaischaemic region of the cardiac muscle in rabbit experiments (104 animals, including 10 controls). A certain periodicity was noted in the changes of the activity of oxidative enzymes in separate zones of the ischaemic myocardium during different periods of the infarction development and healing. The terms of maximum reduction of the enzymatic activity falling within the 4th-6th and 24th hours and 2nd day should be taken into account while developing the methods of metabolic protection and enzyme therapy of the ischaemic myocardium.

Animals↗
PubMedDate not supplied

Interactive static image analysis: a major enhancement to histopathology and cytology.

Recent developments in computerized data processing and digital video technology have made it possible to perform cytology and histopathology in quantitative as well as qualitative terms. The most common method for studying the DNA content of tumors in clinical laboratories is flow cytometry. The development of image analysis (IA) to study DNA content by Feulgen staining and microspectrophotometry actually preceded the development of flow cytometry, but the complexity of the method was so great that it never received widespread usage. In recent years computerized interactive static IA on microscope-based instruments has been developed. These instruments involve microscopy, a television camera and monitor, and a computer to analyze the data. They can be put together from separate components or are available as complete instrument packages from commercial companies. In addition to the study of DNA content, software is also available to perform quantitative immunoperoxidase determination and morphometry. These commercial instruments emphasize user-friendly software. Companies such as CAS (Cell Analysis Systems, Elmhurst, Illinois) provide operator training and annual user meetings. There is a growing interest among laboratories in bringing this technology into their institutions. This review will deal with the following aspects of IA: what it can do, how it works, what skills and training are required for an operator to use it, financial considerations, and how image cytometry compares to flow cytometry.

Cell Biology↗
PubMed1976

[Changes in the activity of 3beta-OH-steroid dehydrogenase in the adrenals of rat fetuses following decapitation in utero].

The activity of 3beta-OH-steroid dehydrogenase in the adrenal cortex of 21-day fetuses in control and 3 days after decapitation in utero was estimated quantitatively by microspectrophotometry. The enzyme activity calculated as per conditioned cell in the fascicular zone exceeded that in the forming glomerular zone. After hypophysectomy the activity of the 3beta-OH-steroid dehydrogenase decreased in both the fascicular and glomerular zones.

Adrenal Cortex↗
PubMedDate not supplied

On the correlation of chromosome number and DNA content in HeLa cell clones.

Chromosome number and nuclear DNA content of two cloned HeLa cell lines were analysed by quantitative Feulgen reaction microspectrophotometry, ultra-rapid microfluorometry and chromosome counting. These cell clones revealed corresponding alterations in their chromosome number and nuclear DNA amount. The results support the hypothesis that a constant relationship exists between both chromosome number and nuclear DNA content in these clonal population.

Cell Count↗