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PubMed1978

Autosomal recessive incomplete achromatopsia with protan luminosity function.

A unique form of dichromatic color vision is described in a family with incomplete achromatopsia. In 1966, incomplete achromatopsia was diagnosed in 4 of 14 children of a consanguineous marriage. The 4 affected had best visual acuities of 6/60 or 6/180, pendular nystagmus, and aversion to bright lights. The ERG showed minimal photopic responses. No abnormality of rod function was present. There was a severe color vision defect. In 1976, one of the patients returned for further color testing. Color tests included measurement of the luminous efficiency function using heterochromatic flicker photometry and colorimetric evaluation. The luminous efficiency function resembled that of the protanope. From the colorimetric measurements, we conclude that the patient has a unique form of dichromatic color vision mediated by two visual photopigments: the normal MWS cone photopigment and a photopigment with the spectral characteristics of rhodopsin.

Adolescent↗
PubMed1995

Decreased dihydropyridine receptor number in hypertensive rat vascular muscle cells.

To further investigate the altered function of Ca2+ channels in vascular muscle cells in hypertension, a novel fluorescently labeled dihydropyridine was used with ultrahigh-sensitivity photometry to study dihydropyridine binding sites on the surface membrane of living vascular muscle cells from stroke-prone spontaneously hypertensive rats and their normotensive controls. Fluorescent nitrobenzoxadiazol-6-dihydropyridine in concentrations of 1 to 100 nmol/L bound specifically to vascular muscle cells' Ca2+ channels, and was displaced by the unlabeled dihydropyridine analogue or nisoldipine (10 mumol/L). Stroke-prone spontaneously hypertensive rat vascular muscle cells showed significantly decreased binding of nitrobenzoxadiazol-6-dihydropyridine compared with normotensive National Institutes of Health rats. Decreased binding of dihydropyridine by vascular muscle cells from stroke-prone spontaneously hypertensive rats (cells that in other studies show increased Ca2+ channel function) indicates a change in channel regulation that is possibly due to a deficiency in the inactivation mechanism, consistent with our earlier electrophysiological studies reporting deficiencies in Ca(2+)-dependent inactivation in genetic hypertension. These data demonstrate decreased numbers of localized sites of dihydropyridine binding on the sarcolemma of living vascular muscle cells, and support the hypothesis that Ca2+ channel alterations may significantly contribute to the molecular etiology of genetic hypertension.

4-Chloro-7-nitrobenzofurazan↗
PubMed1999

Purinoceptor-mediated calcium signaling in preglomerular smooth muscle cells.

-The current studies were performed to determine the contribution of calcium mobilization and voltage-dependent calcium influx to the increase in [Ca2+]i elicited by ATP and UTP. Suspensions of freshly isolated smooth muscle cells were prepared from preglomerular microvessels by enzymatic digestion and loaded with the Ca2+-sensitive dye fura 2. The effect of ATP and UTP on [Ca2+]i was studied on single cells with standard microscope-based fluorescence photometry techniques. Resting [Ca2+]i averaged 80+/-3 nmol/L (n=219 single cells from 58 dispersions). ATP (100 micromol/L) increased [Ca2+]i to a peak value of 845+/-55 nmol/L (n=70 single cells from 38 dispersions) before stabilizing at 124+/-81 nmol/L. Similarly, 100 micromol/L UTP (n=39 single cells from 26 dispersions) stimulated a peak increase in [Ca2+]i of 1426+/-584 nmol/L before reaching a stable plateau of 123+/-10 nmol/L. The [Ca2+]i response to ATP and UTP was also assessed in the absence of extracellular calcium. In these studies, exposure to 100 micromol/L ATP induced a transient peak increase in [Ca2+]i, with the plateau phase being totally abolished. In contrast, exposure to 100 micromol/L UTP under calcium-free conditions resulted in no detectable change in the UTP-mediated increase in [Ca2+]i. The role of L-type calcium channels in the response was assessed with the calcium channel antagonist diltiazem. Incubation with diltiazem (10 micromol/L) markedly reduced the response to ATP, whereas the response to UTP was only slightly reduced. These data demonstrate that both ATP and UTP directly stimulate a biphasic increase in [Ca2+]i in renal microvascular smooth muscle cells. Furthermore, the data suggest that the elevation of [Ca2+]i elicited by ATP is largely dependent on calcium influx through L-type calcium channels, whereas the response to UTP appears to derive primarily from mobilization of calcium from intracellular stores.

Adenosine Triphosphate↗
PubMed1985

Diabetes-induced alterations in the translational activity of specific messenger ribonucleic acids isolated from rat hearts.

During diabetes mellitus, total proteins and ribonucleic acids are significantly decreased in the rat heart, and these parameters can be increased by insulin administration. To determine whether all ribonucleic acids are equally sensitive to insulin, we examined the influence of this hormone on individual translatable ribonucleic acids. Cardiac ribonucleic acid prepared from control, untreated, and insulin-treated diabetic animals was translated in vitro in the presence of [35S]methionine. The radiolabeled peptides were separated by two-dimensional gel electrophoresis and were analyzed by fluorometry. We found that diabetes induces both qualitative and quantitative changes in the predominance of a few specific translatable messenger ribonucleic acid species. The translation of 11 messenger ribonucleic acid species was significantly decreased and that of eight messenger ribonucleic acid species was significantly increased in diabetic preparation. Twelve of the 19 translation products were quantified by digital matrix photometry: three labeled peptides were observed only when cardiac ribonucleic acid from diabetic animals was added to the cell-free translation system, four new peptides appeared when cardiac ribonucleic acid from control animals was added, and although the remaining five peptides were translated in vitro after either control or diabetic ribonucleic acid was added, their relative predominance was altered 2- to 200-fold. When translation products coded for by messenger ribonucleic acids prepared from either diabetic or hypothyroid hearts were compared, we found that most of the alterations induced by diabetes were also induced by hypothyroidism.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Three types of naturally occurring modified lipoproteins induce intracellular lipid accumulation due to lipoprotein aggregation.

Low density lipoprotein (LDL) from patients with coronary atherosclerosis and diabetes mellitus as well as in vitro desialylated LDL, glycosylated LDL, and lipoprotein (a) caused a twofold to fourfold rise in cholesteryl ester in cultured human blood monocytes and intimal smooth muscle cells isolated from normal aorta. Native LDL from healthy subjects failed to induce intracellular lipid accumulation. We have demonstrated by laser correlative photometry and gel filtration chromatography that in vivo and in vitro modified lipoproteins form aggregates under cell culture conditions. The degree of modified lipoprotein aggregation directly correlated with the ability of these lipoproteins to elevate the cholesteryl ester content of cultured cells. Modified lipoprotein aggregates isolated by gel filtration induced a threefold to fivefold elevation in cellular cholesteryl ester content. Aggregates of 125I-modified LDL were taken up and degraded fivefold to sevenfold more effectively as compared with nonaggregated lipoproteins. The uptake and degradation of 125I-labeled aggregates were strongly inhibited by unlabeled aggregates, latex beads, and cytochalasin B but not by native or acetylated LDL. These data indicate that uptake of lipoprotein aggregates occurred by phagocytosis. Obtained results suggest that modified lipoprotein aggregation may be the key condition for lipid accumulation.

Cells, Cultured↗
PubMedDate not supplied

Brain edema during ischemia and after restoration of blood flow. Measurement of water, sodium, potassium content and plasma protein permeability.

The left cerebral hemisphere of Mongolian gerbils was used to elucidate the mechanisms of brain edema which develop during cerebral ischemia and after restoration of cerebral blood flow following temporary ischemia. Water content was measured by the tissue-drying method. Sodium and potssium ion concentration was measured by flame photometry. Passage of 131I-albumin (RISA) from blood to the cerebral parenchyma was measured on a gamma scintillation counter. Our findings indicate that pure cytotoxic edema develops during ischemia and during a short period after restoration of cerebral blood flow. Vasogenic edema, which is accelerated by the leakage of plasma constitutents from blood due to blood-brain barrier damage, developed after restoration of the cerebral blood flow. After less than 1 hr of ischemia, restoration of the cerebral blood flow drastically reduced the degree of brain edema. However, restoration of the cerebral blood flow greatly worsened the brain edema following more than 3 hr of ischemia.

Animals↗
PubMedDate not supplied

Effect of steroid on ischemic brain edema. Analysis of cytotoxic and vasogenic edema occurring during ischemia and after restoration of blood flow.

Mongolian gerbils were observed for the effects of beta-methasone on ischemic brain edema which developed during ischemia or after blood flow restoration. The severity of brain edema was determined by measuring water content of the ischemic cerebral hemisphere, using the wet and dry methods. Sodium and potassium ions were extracted from homogenized brains with 0.75N HNO2 and ion concentration measured by flame photometry. Passage of RISA from blood into the cerebral parenchyma, as an indicator of blood-brain barrier change, was determined with a gamma-scintillation counter. In the cytotoxic edema model, animals were killed after 9 h permanent ischemia or 3 h after 1 h ischemia. In the simultaneous cytotoxic and vasogenic edema model, the animals were killed either 20 h or 3 days following 1 h ischemia, or 3 h after blood flow restoration following 6 h ischemia. Steroid treatment was ineffective in ischemic brain edema of the cytotoxic or vasogenic type.

Animals↗
PubMed1990

Brain edema and cerebrovascular permeability during cerebral ischemia in rats.

Focal cerebral ischemia was produced by occluding the left middle cerebral artery in 769 rats. Permeability of the blood-brain barrier to small or large molecules was evaluated qualitatively using Evans blue or sodium fluorescein and quantitatively using the transfer indexes of iodine-125-labeled bovine serum albumin or [14C]sucrose. Water content was determined using wet and dry weights and sodium and potassium contents using flame photometry. Cortical tissue in the middle cerebral artery territory was sampled less than or equal to 14 days after occlusion. A significant increase in the albumin transfer index was first found 12 hours after occlusion, and the index remained approximately the same until water content peaked 3 days after occlusion. In contrast, the sucrose transfer index increased gradually, significantly correlated with increases in the water and sodium contents. Tissue staining by sodium fluorescein was more extensive than that by Evans blue. As edema fluid decreased gradually 4-10 days after occlusion, the albumin and sucrose transfer indexes increased markedly. These findings indicate that disruption of the blood-brain barrier to small molecules is accompanied by accumulation of edema fluid during the later stages of ischemia. Opening of the barrier to serum protein is probably related to the resolution of edema.

Animals↗
PubMed1991

Intersubject variability in particle deposition does not explain variability in responsiveness to methacholine.

How variable is the deposition of inhaled methacholine (MCH) in the respiratory tract during a challenge test? Does this variability contribute to the variability of airway responsiveness? To examine these questions we estimated the deposition of polydisperse MCH droplets by measuring the deposition of surrogate diethylhexyl sebacate (DEHS) droplets that were similar in size (1.5 microns) but monodisperse. Light scattering photometry and flow measurements were used to compute inspired and expired DEHS particle number. Deposition of DEHS during 4 breaths was measured twice at baseline and after every dose of MCH during an abbreviated challenge test in 16 subjects. Deposition was then compared with reactivity. Reactivity to MCH was expressed as the dose-response slope; it was calculated as percent final change in FEV1/cumulative dose MCH inhaled. Dose-response slopes ranged from zero (nonreactive) to -15.0 (very reactive) %/mumol (mean -3.2 +/- 5.3 SD). Seven subjects had a 20% or greater decrement in FEV1 after their highest MCH dose. Baseline DEHS deposition, which ranged from 66 to 84% (mean 77 +/- 5 SD), was not significantly different between responders and nonresponders and was not a significant predictor of the dose-response slope. Reactivity was significantly associated with an increase in deposition produced by MCH (p less than 0.007). This increase was small, however (relative change less than 7%), so that the effect on the deposited dose of MCH was minimal. We conclude that, with the breathing pattern used, individual differences in DEHS (and MCH) deposition were small and contributed little to intersubject variability of responsiveness to inhaled MCH.

Adult↗
PubMed1990

The effects of activated eosinophils and neutrophils on guinea pig airway epithelium in vitro.

Epithelial shedding is a characteristic feature of asthmatic airways and has been attributed to eosinophil products. We have examined the interaction of purified intraperitoneal guinea pig eosinophils with or without platelet-activating factor (PAF, 10(-7) M) or lyso-PAF (10(-7) M) with guinea pig tracheal epithelium in vitro. At 0, 4, 14, and 24 h, the percentage of ciliation of the tracheal circumference (CTC) was measured by light microscopy and the ciliary beat frequency (CBF) by photometry. PAF-activated eosinophils (50 x 10(6) cells/ml) disrupted the epithelium, mean CBF and CTC being reduced by 77.8 +/- 5.8% (mean +/- SEM; P less than 0.001 versus control) and 94.2 +/- 1.4% (P less than 0.001) over 24 h, respectively. PAF (10(-7) M) alone had no significant effect. Lyso-PAF with eosinophils (50 x 10(6) cells/ml) also reduced mean CBF and CTC but to a lesser extent. Eosinophils alone also led to a reduction of 36.2 +/- 11.4% in mean CBF and 53.0 +/- 15.5% in CTC, but these changes were not significant. The PAF antagonist, WEB 2086 (10(-6) M), significantly inhibited the mean CBF and CTC reduction due to PAF-activated eosinophils by 61.5 +/- 17.2% (P less than 0.01) and 20.8 +/- 6.5% (P less than 0.05), respectively. In addition, catalase (1,125 U/ml) partially inhibited the mean CBF and CTC reduction induced by PAF-activated eosinophils. Intraperitoneal neutrophils (PMN) (50 x 10(6) cells/ml) also disrupted epithelium but to a lesser extent (24-h reduction: 34.2 +/- 12.7% for mean CBF and 60.2 +/- 13.2% for CTC, respectively). Stimulation with PAF (10(-7) M) had no further effect. Marked exfoliation of the epithelial layer was observed after 14 h of incubation with activated eosinophils. We concluded the PAF-activated eosinophils are capable of grossly disrupting ciliated epithelium and may contribute to epithelial damage observed in asthma.

Animals↗
PubMed2001

Photopigment basis for dichromatic color vision in the horse.

Horses, like other ungulates, are active in the day, at dusk, dawn, and night; and, they have eyes designed to have both high sensitivity for vision in dim light and good visual acuity under higher light levels (Walls, 1942). Typically, daytime activity is associated with the presence of multiple cone classes and color-vision capacity (Jacobs, 1993). Previous studies in other ungulates, such as pigs, goats, cows, sheep and deer, have shown that they have two spectrally different cone types, and hence, at least the photopigment basis for dichromatic color vision (Neitz & Jacobs, 1989; Jacobs, Deegan II, Neitz, Murphy, Miller, & Marchinton, 1994; Jacobs, Deegan II, & Neitz, 1998). Here, electroretinogram flicker photometry was used to measure the spectral sensitivities of the cones in the domestic horse (Equus caballus). Two distinct spectral mechanisms were identified and are consistent with the presence of a short-wavelength-sensitive (S) and a middle-to-long-wavelength-sensitive (M/L) cone. The spectral sensitivity of the S cone was estimated to have a peak of 428 nm, while the M/L cone had a peak of 539 nm. These two cone types would provide the basis for dichromatic color vision consistent with recent results from behavioral testing of horses (Macuda & Timney, 1999; Macuda & Timney, 2000; Timney & Macuda, 2001). The spectral peak of the M/L cone photopigment measured here, in vivo, is similar to that obtained when the gene was sequenced, cloned, and expressed in vitro (Yokoyama & Radlwimmer, 1999). Of the ungulates that have been studied to date, all have the photopigment basis for dichromatic color vision; however, they differ considerably from one another in the spectral tuning of their cone pigments. These differences may represent adaptations to the different visual requirements of different species.

Adaptation, Physiological↗
PubMed2003

Light exposure and macular pigment optical density.

PURPOSE: Biochemical research has demonstrated that lutein and zeaxanthin, the two macular carotenoids, are bleachable pigments. Further, evidence suggests that exposure to UV light can degrade plasma carotenoid levels in vivo. The present study investigated the effects of exposure to normal levels of light on the levels of lutein and zeaxanthin in the retina. METHODS: The optical density of macular pigment (MPOD) was measured in two male subjects under four different light-adaptation conditions for 20 days. Heterochromatic flicker photometry was used to measure MPOD at 0.5 degrees eccentricity. RESULTS: The four conditions of light adaptation did not significantly affect MPOD. As in previous studies, however, a significant day-to-day difference was observed for both subjects. CONCLUSIONS: The results suggest that lutein and zeaxanthin levels in the eye are unaffected by light and oxidation throughout the day. This justifies current research methods in which MPOD measures are made regardless of the time of day. However, significant between-day variance indicates that multiple MPOD measures may be necessary to evaluate lutein and zeaxanthin levels in the retina accurately.

Adaptation, Ocular↗
PubMed2003

Increased endothelial survival of organ-cultured corneas stored in FGF-2-supplemented serum-free medium.

PURPOSE: To investigate the effect of FGF-2 on corneal endothelial cell survival in porcine and human corneas during corneal storage in a serum-free medium. METHODS: Porcine and paired human corneas were stored at 32 degrees C for 9 and 22 days, respectively. One cornea of each pair was stored in a serum-free culture medium, and the mate was preserved in the same medium supplemented with 10 ng/mL FGF-2. Quantitative analysis of corneal damage after storage was determined by the Janus green photometry technique. 5-Bromo-2-deoxyuridine (BrdU) labeling of the endothelium determined the effect of FGF-2 on endothelial proliferation during storage. Additional cell culture studies were performed to elucidate the role of FGF-2 on the incidence of endothelial apoptosis after serum deprivation. RESULTS: When FGF-2 was added to the serum-free medium, the damage rates of porcine endothelia were reduced from 15.1% +/- 8.7% (control) to 6.4% +/- 2.0% after 9 days and from 25.3% +/- 10.2% to 13.6% +/- 4.2% after 22 days of storage. In the human corneas stored during 22 days in FGF-2-supplemented medium, the amount of endothelial damage was 11.8% +/- 3.2%, which was significantly less damage than in the control fellow corneas stored in unsupplemented serum-free medium (19.3% +/- 6.3%; P < 0.01). DNA synthesis was not enhanced in corneas stored in serum-free medium, serum-free medium+FGF-2, or medium containing 10% FCS. Only a few (3.8%) TUNEL-positive endothelial cells were detected in cultures maintained in FGF-2-supplemented serum-free medium compared with a high number (48%) of apoptotic cells in control cultures. CONCLUSIONS: FGF-2 efficiently reduces human corneal endothelial damage that occurs during organ culture storage in a serum-free medium. This effect is truly protective, because no proliferative activity and a decreased rate of apoptosis were determined. FGF-2 emerges as an important component of a future serum-free corneal organ-culture medium established to replace fetal calf serum (FCS) as a potential source of recipient infection.

Aged↗
PubMed2004

Spatial properties of photophobia.

PURPOSE: To determine the spatial properties of stimuli that elicit photophobia (PP) in normal subjects: Does PP exhibit spatial summation? Are different parafoveal quadrants (superior, inferior, temporal, and nasal) of the retina differentially sensitive in PP? What is the relationship between PP sensitivity and retinal eccentricity? What is the relationship between the spatial properties of PP and the spatial distribution of macular pigment (MP)? METHODS: A Maxwellian-view optical system with a xenon light source was used to present the stimuli. Four normal subjects viewed stimuli of various sizes, retinal locations, and one of two chromatic contents: xenon-white and a broadband orange. The intensity of the test stimulus was increased between trials until the PP threshold was reached. The squinting response corresponding to PP was assessed by electromyography and used as an objective criterion of PP. Three parameters were examined: stimulus size, parafoveal retinal locus (superior, inferior, temporal, and nasal), and retinal eccentricity (extending into the perifovea). Spatial profiles of MP were measured psychophysically using heterochromatic flicker photometry (HFP). RESULTS: Spatial summation for PP was found essentially to adhere to Piper's law (radiance proportional to square root of stimulus area). The PP response was greater to centrally than peripherally viewed targets. In this regard, MP acted as a spatially integrated filter in the attenuation of PP. CONCLUSIONS: The degree of spatial summation found for PP indicates that an increase of 1.0 log unit in field area results in an approximately 0.57-log-unit decrease in the radiance required to elicit PP. PP appears to serve the function of retinal photoprotection.

Adult↗
PubMed1991

Development and evaluation of the simultaneous determination of sweat sodium and chloride by ion-selective electrodes.

A method for the simultaneous determination of sweat sodium and chloride was developed and evaluated using a Monarch 2000 centrifugal analyser with ion-selective electrodes. In order to measure sodium and chloride in diluted sweat samples and sweat paper eluates an analytical working range of below 10 mmol/L was required. This was achieved by modification of the Monarch ISE software parameters for urine electrolytes together with the use of calibrators within this range. Evaluation of the method has included linearity, recovery and precision studies and the correlation between ion-selective electrode and flame photometry for sweat sodium and between ion-selective electrode and a continuous flow method for sweat chloride. Of the patients studied so far, there has been good discrimination in sweat electrolytes between those with and without cystic fibrosis.

Centrifugation↗
PubMed1994

The analysis of metabolites in human sweat: analytical methods and potential application to investigation of pressure ischaemia of soft tissues.

A straightforward technique was developed for sweat collection applicable to tissues subjected to external load without introducing distortion of underlying tissues, and for analysis of six metabolites in the collected sweat. Chloride was measured colorimetrically and lactate, urea and urate by enzymatic methods on a centrifugal analyser. Sodium and potassium were measured by flame photometry. The methods showed good precision, recovery and linearity. To assess the technique sweat was collected: (i) from the sacrum, ischium, forearm and calf in healthy individuals at 32 degrees C for 1 h; (ii) from the sacrum of healthy subjects at ambient temperature for 9 h; (iii) at ambient temperature from the sacrum of a patient with a history of pressure sores. Sweat rates were greater at the sacrum and ischium than the calf or forearm. There were differences in the concentrations of lactate and urea between sites but these were smaller when expressed as amount secreted. Sweat rates were significantly lower in groups (ii) and (iii), but sweat could be collected reliably. This technique has potential clinical application to the investigation of susceptibility to pressure sores.

Adult↗
PubMed2002

DNA extraction from archival formalin-fixed, paraffin-embedded tissue sections based on the antigen retrieval principle: heating under the influence of pH.

During the course of diagnostic surgical pathology, pathologists have established a large collection of formalin-fixed, paraffin-embedded tissues that form invaluable resources for translational studies of cancer and a variety of other diseases. Accessibility of macromolecules in the fixed tissue specimens is a critical issue as exemplified by heat-induced antigen retrieval (AR) immunohistochemical (IHC) staining. On the basis of observations that heating may also enhance in situ hybridization (ISH) and the similarity of formalin-induced chemical modifications that occur in protein and in DNA, we designed a study to examine the efficiency of DNA extraction from archival formalin-fixed, paraffin-embedded tissues using an adaptation of the basic principles of the AR technique, i.e., heating the tissue under the influence of different pH values. Archival paraffin blocks of lymph nodes, tonsil, and colon were randomly selected. Each paraffin block was prepared in 34 microtubes. For each paraffin block, one tube was used as a control sample, using a non-heating DNA extraction protocol. The other 33 tubes were tested using a heating protocol under 11 variable pH values (pH 2 to 12) under three different heating conditions (80, 100, and 120C). Evaluation of the results of DNA extraction was carried out by measuring yields by photometry and PCR amplification, as well as kinetic thermocycling (KTC)-PCR methods. In general, lower pH (acid) solutions gave inferior results to solutions at higher pH (alkaline). Heating tissues at a higher temperature and at pH 6-9 gave higher yields of DNA. There appeared to be a peak in terms of highest efficiency of extracted DNA at around pH 9. The average ratios 260:280 of extracted DNA also showed better values for samples heated at 120C. PCR products of three primers showed satisfactory results for DNA extracted from archival paraffin-embedded tissues by heating protocols at pH 6-12, with results that were comparable to the control sample subjected to the standard non-heating, enzymatic DNA extraction method. This study is the first to document the use of heating at an alkaline pH for DNA extraction from archival formalin-fixed, paraffin-embedded tissues, a recommendation based on the principles of AR for protein IHC. These findings may lead to a more effective protocol for DNA extraction from archival paraffin-embedded tissues and may also provide enhanced understanding of changes that occur during formalin-induced modification of nucleic acids.

Antigens↗
PubMed1986

Quantitative histochemistry of benzaldehyde dehydrogenase in hepatocellular carcinomas of vinyl chloride-treated rats.

Hepatocarcinogenesis in rats treated with several chemicals is associated with changes in aldehyde dehydrogenase (AlDH) activity, particularly heterogeneous expression of a "tumor specific" phenotype that is very active with aromatic aldehydes, e.g., benzaldehyde (Bz). Objectives of this study were first, to determine if liver cancers in vinyl chloride-treated rats also expressed this AlDH phenotype, and second, to quantitate the NAD- and NADP-dependent AlDH activity for the substrates Bz and acetaldehyde (Ac) in the cancers and surrounding tissue. Small cubes of tissue containing well-differentiated hepatocellular carcinoma were obtained from five Sprague-Dawley rats exposed to 2500 ppm vinyl chloride for 55 weeks. An optimized procedure was developed for AlDH histochemistry. Frozen sections were preincubated in nitroblue tetrazolium/acetone and then incubated at 20 degrees C in viscous polyvinyl alcohol media containing buffer, phenazine methosulfate, sodium azide, substrate, coenzyme, and nitroblue tetrazolium. Background activity was evaluated by omission of substrate. Activity was quantitated by computer-assisted microscopic photometry. All five carcinomas had heterogeneous staining of NADP- and NAD-dependent BzDH and AcDH activity, with clusters of very high-activity cells. The magnitude of staining in the high-activity neoplastic cells was at least tenfold greater for BzDH-NADP and about twofold greater for BzDH-NAD, AcDH-NADP, and AcDH-NAD than the staining in other liver cells. More neoplastic cells had high BzDH than high AcDH activity. Only BzDH-NADP was localized predominantly to the carcinoma.

Aldehyde Dehydrogenase↗