Occurrence of lipids in the liver of the hypertriglyceridemic rats.

AIMS: The purpose of this study was to demonstrate the accumulation and distribution of lipids in the liver of the adult Prague hereditary hypertriglyceridemic (HHTg) rats. They reveal an increased expression of 11beta-hydroxysteroid dehydrogenase 1 (11HSD1), which locally increases concentration of corticosterone in the liver. We studied the effect of the 11HSD1 inhibition on the lipid content. METHODS: Samples of liver of three groups of adult female rats--HHTg, HHTg treated for 14 days with 50 mg/kg/day carbenoxolone (HHTg+CBX) and control Wistar rats, were examined histochemically. Cryosections of the samples were stained with Oil red O or Sudan black B to demonstrate different kinds of lipids. Extent and intensity of staining was evaluated semiquantitatively. RESULTS: The orientational analysis showed a higher extent and intensity of the staining of the liver of HHTg and HHTg+CBX rats (equal in both hypertriglyceridemic groups) than that of the control Wistar rats. Oil red O stained unsaturated fatty acids and neutral fats, mainly triglycerides. The difference was on average 30 per cent. Staining of phospholipids with Sudan black B showed similarly the higher positivity in the hypertriglyceridemic groups than in controls. CONCLUSIONS: Staining for triglycerides and phospholipids demonstrated a higher amount of lipids in the liver of HHTg and HHTg+CBX female rats than in controls. The inhibition of 11HSD1 activity had no effect on the lipid content in the liver of the HHTg rats.

Determination of uric acid in plasma and allantoic fluid of chicken embryos by capillary electrophoresis.

Capillary electrophoresis with diode array detection (DAD) was used to determine uric acid (UA) in chicken plasma and the allantoic fluid of chicken embryos. Complete separation of uric and ascorbic acids was attained in less than 10 min in the optimized BGE containing 60 mM MES + 30 mM Tris + 0.001% (w/v) polybrene (pH 6.1). The limit of UA detection (0.2 mg/L) was found to be low enough for sensitive analysis of native plasma and allantoic fluid samples. Range of linearity (1-200 mg/L), repeatability for peak area (CV <4.1%) and migration time (CV < 2.5%), as well as recovery of UA from biological samples (97-100%), were found to be satisfactory. The method was applied to detect the elevated UA concentrations (hyperuricemia) in chicken embryos with induced unilateral renal agenesis. CE/DAD analysis of the chicken plasma can be carried out with a relatively small volume of samples (1 microL).

Indicators of functional differentiation of the chick embryonic kidney.

Relevant indicators of the functional capability of the embryonic kidney were tested in the chick mesonephros chosen as an ideal model accessible to direct observation in vivo. Evidence of glomerular filtration (GF) was checked up by the arterial injection of 2% lissamine green (LG) followed by measurement of the LG passage time on days 5, 6 and 7. Presence of the electrogenic transport was investigated by determining the transepithelial potential difference (TPD) which distinguished proximal and distal tubules of the 6-day nephrons. GF and tubular reabsorption could be demonstrated from day 5 by the storage of trypan blue (TB) in proximal tubules after intra-amniotic administration of the dye. The distribution of tubule staining corresponded to the proximal-distal gradient of the nephron differentiation. Activities of embrane enzymes, alkaline phosphatase and 5'-nucleotidase, were detected from day 4. They preceded the ultrastructural maturation in the differentiating proximal tubule epithelia. A semiquantitative evaluation of enzyme activities by the method of measuring of the minimum incubation time (MIT) together with the TB storage, appeared reliable and relevant indicators of the functional properties of mesonephric nephrons, suitable for distinguishing between more and less advanced stages of the nephron development.

Apical ouabain-sensitive and ouabain-insensitive ATPases in rat colonic epithelium.

Active resorption of potassium ions in the colon is mediated by ouabain-sensitive and ouabain-insensitive H,K-ATPases localized in the apical membrane of colonic enterocytes (colonocytes). The present study was performed to investigate distribution patterns of apical ATPases using catalytic histochemistry and immunohistochemistry. Activity of the ATPases was localized both at the apical and basolateral regions of these cells. The basolateral activity was almost completely inhibited by ouabain, but apical ATPase activity was only partially inhibited by ouabain. Ultracytochemically, activity was localized at the cytoplasmic side of the apical and basolateral membrane and thus we assume that the activity represents isoforms of apical H,K-ATPases and basolateral Na,K-ATPase. With the use of a polyclonal antibody raised against H,K-ATPase alpha-subunit, we demonstrated immunostaining only in the apical region of colonic enterocytes whereas positive staining was not observed at the basolateral membrane. On the other hand, when antibodies against alpha1-subunit Na,K-ATPase were applied, immunostaining was localized only in the basolateral membrane domain. Therefore, we conclude that ATPases as demonstrated histochemically in the present study were identified immunohistochemically as colonic alpha-subunit H,K-ATPase (in the apical cell membrane of colonocytes along the entire length of crypts) and as alpha1-subunit Na,K-ATPase (in the basolateral membrane).

Histochemical demonstration of the heterogeneity of the epithelium of proximal tubules in the chick mesonephros.

Proximal tubules (PT) in 7-10-day old chick mesonephros were examined histochemically to evaluate their structural and functional properties related to the absorption capacity of the epithelium and its possible alterations leading to cystically dilated tubules (CDT). Alkaline phosphatase activity at the apical cell membrane was demonstrated in varying intensities in large PT. A similar heterogeneity was also detected in the expression of proteoglycans (Lewis(x) antigen) localized in the apical part of the epithelial cell membrane but not in the basolateral membrane parts (beta-catenin, Na(+),K(+)-ATPase). In analogy, the ability to accumulate trypan blue was found in the same cell population. We hypothesize that epithelial cells in proximal tubules of nephrons with a defective apical cell membrane cause reduced fluid absorption and subsequent overfilling and dilation of the tubules.