Localization of thyroxine 5'-monodeiodinase activity in the renal proximal tubules in rabbits and rats

To determine the localization of T4 5'-monodeiodinase activity in rabbit and rat nephron segments, the formation of tri-iodothyronine (T3) from thyroxine (T4) was measured in kidney homogenate and in isolated nephron segments obtained by the microdissection method. In order of decreasing activity, homogenates of rabbit renal cortex, outer medulla and inner medulla were capable of converting T4 to T3. In the isolated nephron segments of the rabbit cortex, the activities were noted in both proximal convoluted and proximal straight tubules. On the other hand, the activities were not detected in segments including the cortical thick ascending limb of Henle's loop, the distal convoluted tubule, the connecting tubule, and the cortical collecting tubule. It is concluded that both the convoluted and the straight tubules are the sites of T3 production in the kidney.     

Thyroxine 5'-monodeiodinase activity in regenerating liver of triiodothyronine-treated rats

The effect of triiodothyronine (T3) on hepatic thyroxine (T4) 5'-monodeiodinase and the subcellular localization of the enzyme were examined in regenerating rat liver, because it seemed likely that the effect of T3 might be accentuated during liver regeneration. Five days after T3 treatment, the specific activity of the monodeiodinase in the microsomal fraction (105,000 X g pellet) of regenerating liver was increased to 207% of the control value. Lineweaver-Burk plots showed that the Vmax for T4 5'-monodeiodination was about 3 times greater in T3-treated rats than in controls, but that there was no difference between the two groups in the apparent Km value for T4. About 55% of the total enzyme activity was found in the endoplasmic reticulum (ER) of the liver of both controls and T3-treated rats. The subcellular distribution of the enzyme was similar to that of NADPH-cytochrome c reductase (NADPH-cyt c reductase), a marker of the ER, but different from that of Na+,K+-ATPase, a marker of plasma membranes (PM).     

Subcellular localization of diamine oxidase in rabbit kidney cortex

The intracellular localization of diamine oxidase (EC 1.4.3.6) in rabbit kidney cortex was studied. The distribution of diamine oxidase in the subcellular fractions, obtained by modifying the classical method of Wattiaux-De Coninck, S., Rutgeerts, M.T. and Wattiaux, R. (Biochim. Biophys. Acta (1965) 105, 446-459) demonstrated that this activity is concentrated (greater than 60%) in the microsomal fraction. Biochemical and morphological data indicate a 20-30% contamination of this fraction by plasma membrane and brush border fragments. Subfractionation of the microsomes, obtained by centrifuging in a continuous sucrose-Ficoll gradient (d 1.038-1.064) for 75 min, showed that diamine oxidase is concentrated in membrane deriving from the endoplasmic reticulum. In fact the bulk of diamine oxidase activity was recovered in a subfraction of the gradient which was shown both biochemically and morphologically to derive from the endoplasmic reticulum. The possible significance of this result is discussed.     

Growth hormone stimulates the peripheral conversion of thyroxine into triiodothyronine by increasing the liver 5'-monodeiodinase activity in the fasted and normal fed chicken

Normal fed and 2 days fasted Warren chickens were injected intravenously with 100 micrograms of ovine growth hormone (GH) and ovine prolactin and plasma concentrations of thyroid hormones were assayed prior and up to 2 h after injection. Fasting alone decreases T3, but increases T4. An injection of GH resulted in increases of plasma T3 concentrations in two fasting experiments by 40% (after 3/4 h) and 104% (after 1 h). In normal fed animals no increase is observed in the first experiment, whereas a 35% increase occurs in the second one. An injection of 100 micrograms prolactin does not influence T3 in normal fed or fasting animals. Both GH and prolactin, however, may decrease plasma concentrations of T4. In a separate experiment 50 micrograms and 200 micrograms of GH raised the decreased T3 levels after fasting by 39% and 60% respectively 1 h after injection and by 24 and 61% respectively in normal fed chicken, whereas prolactin was ineffective in this regard. Using Hisex chickens, the influence of an injection of 100 micrograms GH on plasma concentrations of thyroid hormones could be confirmed. At the same time GH increases the liver 5'-monodeiodinase activity by 330% after 1 h and by 147% after 2 h. The peroxidase activity is not influenced in normal fed chickens, but GH decreases this activity in food deprived animals after 1 h and 2 h. It is concluded that ovine GH, but not prolactin, stimulates the peripheral conversion of T4 into T3 in both normal fed and food deprived chicken and that this effect is dose dependent.     

Investigation of thyroxine monodeiodinase activity in the liver, kidney and brown adipose tissue of foetal and neonatal rabbit

The maturation of the 5'- and 5-monodeiodinase system in liver, kidney and brown adipose tissue of rabbits, during the foetal period (from 21 days of gestation to birth) and the neonatal period (from birth to 3 weeks of life) was studied. A sudden increase of 5'- and 5-monodeiodinase activity in liver and kidney 3 days before birth was observed, falling to a nadir at day 3 after birth. Foetal and neonatal serum T4, T3 and rT3 concentration were very low and rose progressively with age, reaching adult values at about day 21. In the foetal brown adipose tissue high 5'-monodeiodinase and low 5-monodeiodinase activity was found. The 5'-monodeiodinase decreased during the first days of life whereas the 5-monodeiodinase activity remained at a low stable level until day 3 when the activities of both enzymes increased. The increase of conversion rate of T4 to T3 and rT3 in liver and kidney well correlate with the triiodothyronines concentration in serum from day 3 after birth.     

Subcellular distribution of hypotaurine oxidase activity in ox retina

Hypotaurine oxidase activity was determined in the primary and secondary subfractions of ox retina. About 78% of enzyme activity was found in the soluble fraction while about 22% was associated with particulate components.In the secondary subcellular fractions about 55% of enzyme activity, recovered from crude mitochondria, was present in the synaptosomal fraction.This finding suggests that the conversion of hypotaurine to taurine is possible in ox retina.     

Subcellular distribution and properties of kynurenine pyruvate transaminase in rat kidney.

Kynurenine transaminase activity in rat kidney was found in both the mitochondrial and supernatant fractions. These fractions contained (a) kynurenine pyruvate transaminase, which showed a preference for pyruvate as amino acceptor, and had a pH optimum between 8.0 and 8.5, and (b) kynurenine 2-oxoglutarate transaminase, with a preference for 2-oxoglutarate and a pH optimum between 6.0 and 6.5. The apparent Km value of the former enzyme for L-kynurenine was much lower than that of the latter enzyme.     

Different responses between thyroxine 5'-monodeiodinase and protein disulfide isomerase activities to dietary fat and thyroid status in rat liver

On the hypothesis that rat hepatic microsomal type I iodothyronine 5'-monodeiodinase (MDI) would be identical to hepatic protein disulfide isomerase (PDI) (Boado et al. Biochem. Biophys. Res. Commun. 155 1297-1304 (1988)), we examined the responses of these enzyme activities to dietary fat and thyroid status in rats. The hepatic MDI activity was higher in rats fed high fat diet than in rats fed low fat diet, but the hepatic PDI activity showed the reverse responses to the diets. Propylthiouracil administration (hypothyroidism) lowered the MDI activity, but elevated the PDI activity. Thyroxine administration (hyperthyroidism) elevated the MDI activity but had no effect on the PDI activity. These results indicate that the two enzyme activities are regulated by different mechanisms in vivo, suggesting that MDI and PDI are not identical enzymes.     

Subcellular distribution of cysteine oxidase activity in ox retina.

Cysteine oxidase activity has been determined in the primary and secondary subfractions of ox retina. About 30% of enzyme activity is found in the soluble fraction while about 70% is associated with particulate components.In the secondary subcellular fractions about 36% of enzyme activity, recovered from crude mitochondria, is present in the synaptosomal fraction.Enzymic activity is stimulated by Fe⁺⁺ and NAD⁺. The reason and significance of the cysteine oxidase activity in synaptosomal fraction are briefly discussed in relation with taurine function in retina.     

Serum concentrations of thyroid hormones and extrathyroidal thyroxine-5'-monodeiodinase activity during lactation in the rat

Serum thyroxine (T4) and triiodothyronine (T3) concentrations and T4-5'-monodeiodinase activity in liver and kidney homogenates were studied in Sprague-Dawley rats during lactation. Blood and tissue samples were collected from nulliparous and pregnant rats 2 days before delivery and from lactating rats 0, 2, 7, 12, 19, and 26 days after delivery. Litters were removed from half of the mothers immediately after delivery to create a postpartum nonlactating group for study at the same times. Pregnant rats had lower serum T4 and T3 concentrations and higher liver T4-5'-monodeiodinase activity than nulliparous females. Low serum T4 persisted throughout lactation but further decrease in serum T3 was observed. Activity of T4-5'-monodeiodinase in liver and kidney homogenates was significantly reduced during lactation as compared to nonlactating rats. Serum concentration of T4 and T3 and T4-5'-monodeiodinase activity in liver and kidney returned toward control values 5 days after weaning (Postpartum Day 26). Our findings suggest that the relative hypothyroid state observed during lactation in rats is associated with a significant decrease in T4 to T3 conversion in the liver and kidneys.