Synthesis of a shortened pro-alpha 2(I) chain and decreased synthesis of pro-alpha 2(I) chains in a proband with osteogenesis imperfecta

Synthesis of type I procollagen was examined in skin fibroblasts from a proband with a lethal variant of osteogenesis imperfecta. The fibroblasts synthesized shortened pro-alpha 2(I) chains and these shortened chains accounted for all the pro-alpha 2(I) chains synthesized by the cells. In addition, there was a decrease in the relative rate of synthesis of pro-alpha 2(I) chains. Fragmentation of the shortened pro-alpha 2(I) chains with vertebrate collagenase and cyanogen bromide demonstrated that the shortening was in alpha 2(I)-CB3,5A, a fragment from about the middle of the chain containing amino acid residues 361 to 775. Based on the relative mobility in electrophoretic gels, the shortening was about 20 amino acid residues. The decreased synthesis of pro-alpha 2(I) chains was demonstrated by an increase in the ratio for the rates of synthesis of pro-alpha 1(I):pro-alpha 2(I) chains. It was associated with an increase in the ratio of mRNAs for pro-alpha 1(I):pro-alpha 2(I) in the cells. Fibroblasts from the father also demonstrated a decreased synthesis of pro-alpha 2(I) chains as reflected by an increase in the ratio of newly synthesized pro-alpha 1(I):pro-alpha 2(I) chains. No shortened pro-alpha 2(I) chains were seen in fibroblasts from either the father or the mother. The observations suggested that the proband inherited a nonfunctioning pro-alpha 2(I) gene from her father and that the gene for the shortened pro-alpha 2(I) chain probably arose from a sporadic mutation.     

A dynamometer for the measurement of the extension torque of the lower leg during static and dynamic contractions of the quadriceps femoris muscle

A dynamometer which makes an angular movement is described. The dynamometer enables the measurement of the extension torque of the lower leg at different knee angles during static and slow concentric and eccentric contractions of the quadriceps femoris muscle. The influence of gravity on the measured torque signal can be compensated for by another signal representing the angular movement. The application of the dynamometer is demonstrated by giving an example of measurement.     

Effect of oestradiol and tamoxifen on the testosterone response in male rats to a single injection of hCG

A single s.c. injection of hCG (100 i.u.) produced a biphasic serum testosterone response in adult male rats, peaks being noted at 2 h (24 ng/ml) and 3 days (16 ng/ml). The levels fell to control during the intervening interval (8 ng/ml), although there were elevated levels of serum hCG. Maintenance of high oestradiol levels by a s.c. injection of 50 micrograms oestradiol benzoate given on Day 2 after the initial hCG injection failed to prolong the refractory period and the secondary peak of testosterone (16 ng/ml) occurred on Day 3. Administration of the antioestrogen, tamoxifen (2 mg or 3 micrograms), 24 h before or simultaneously with hCG did not prevent testicular refractoriness in vivo because serum testosterone levels still declined after 2 h to reach a nadir at 2 days. The basal in-vitro testosterone production by decapsulated testes from animals injected with hCG was enhanced at 2 h. Stimulation by hCG increased the amount of testosterone produced (X 1.5 that in controls). By 12 h basal production decreased and there was no further increment in testosterone in the presence of hCG. This refractoriness to further hCG stimulation prevailed until Day 3, but the total production of testosterone fell so that at 24 h and 2 days testes were producing basal amounts of testosterone. Testes recovered from refractoriness at 4 and 5 days, when basal and stimulated testosterone production were greater than in controls. Injection of 50 micrograms oestradiol benzoate at 2 days did not prolong the in-vitro refractory period and 2 mg or 3 micrograms tamoxifen had no effect on the in-vitro steroidogenic activity, since testes were still refractory to further hCG stimulation from 12 h to 3 days. The results of the present study do not support the hypothesis that oestradiol is involved in the hCG-induced refractoriness of the Leydig cell. The nadir between the peaks of serum testosterone in vivo corresponds to the period during which the testis is refractory to in-vitro stimulation by hCG.     

Accumulation of phosphatidic acid in microsomes from propranolol-treated retinas during short-term incubations

Abstract: The pool size and synthesis of phosphatidic acid derived from [2-³H]glycerol were studied in bovine whole retinas and subcellular fractions. Microsomal preparations from retinas incubated with [2-³H]glycerol displayed the highest percentage labeling of phosphatidic acid at 5 min of incubation; labeling decreased rapidly thereafter. In drug-treated retinas,0.5 m M propranolol increased the endogenous content of phosphatidic acid and stimulated [2-³H]glycerol labeling in whole retina and microsomal and postmicrosomal supernatant fractions. This effect was observed during short-term incubations and was reversible. In pulse-chase experiments, 60 min of reincubation greatly reduced the labeling effect, although propranolol still enhanced phosphatidic acid labeling. At the same time, endogenous phosphatidic acid accumulated and reincubation without propranolol reversed the effect. During accumulation, the amount of palmitate increased and that of oleate decreased, whereas the relatively high level of docosahexaenoate in phosphatidic acid remained unchanged. It was concluded that this propranolol-induced effect is due to cationic amphiphilic drug activity in the endoplasmic reticulum that results in a partial inhibition of phosphatidic acid degradation and a stimulation of its de novo synthesis. Hence, net synthesis of phosphatidic acid can be assessed in the retina during short-term incubation with propranolol.     

Trypanosoma cruzi: distribution of fluorescently labeled tubulin and actin in epimastigotes

Cytoskeletal components were visualized in epimastigote forms of Trypanosoma cruzi by double immunofluorescence microscopy using monospecific antibodies against tubulin and against actin. Intense staining of the flagellum and the edges of the cell body was observed when the cells were stained with anti-tubulin, reflecting the presence of the basal bodies, the flagellar axoneme and the subpellicular microtubules. A less intense staining was seen in the cell body of epimastigotes stained with anti-actin. However, an intense staining was observed with this antibody in the flagellum, in a pattern similar to that observed with anti-tubulin. It is suggested that the paraxial structure, which is formed by a complex array of 6-nm-thick microfilaments is composed, at least in part, of actin.     

Arginine catabolism in the cotyledons of developing and germinating pea seeds

Arginine is the predominant free amino acid in the cotyledons of developing seeds of Pisum sativum L. cv Marzia. Breakdown of arginine was measured by injecting l-[guanido-¹⁴C]arginine into detached cotyledons. Cotyledons of developing seeds showed a low rate of ¹⁴CO₂ evolution whereas a much higher rate of ¹⁴CO₂ evolution was measured from cotyledons of seeds 4 days after the onset of germination. The activities of the catabolic enzymes arginase, urease, and ornithine aminotransferase were measured throughout development and germination. Arginase and ornithine aminotransferase were present at an early stage of development. Urease activity appeared later as the seeds started to desiccate. During germination, all three enzymes were present. The different course of activity of these enzymes indicates that they are controlled separately.     

Biochemical parameters to assess cell differentiation of Bouvardia ternifolia Schlecht callus

Calli derived from leaves and radicles of B. ternifolia were grown on Murashige and Skoog (MS) basal medium, and the effects of different nitrogen sources on the rate of callus growth and on the enzymes related to nitrogen assimilation were studied. Ammonium alone did not support callus growth unless a Krebs-cycle intermediate was added to the medium. The activities of glutamine synthetase (EC 6.3.1.2), glutamate synthase (EC 1.4.7.1), and glutamate dehydrogenase (EC 1.4.1.2) were measured in homogenates of callus grown on media supplied with different nitrogen sources. The results indicate that leaf and root calli have similar levels of these enzymes when grown on MS medium (Murashige and Skoog 1962. Physiol. Plant. 15, 473–497). However, when the calli were supplied with glutamine as the sole nitrogen source, the activity of glutamate synthase increased in leaf callus but was almost completely inhibited in root callus. The results indicate that calli originated from different B. ternifolia tissues do not have the same biochemical dedifferentiated state.     

Long-acting contraceptive agents: norethisterone esters of monoalkenyl and monoalkynyl acids

The synthesis of nine new esters of norethisterone (17 alpha-ethynyl-17 beta-hydroxyestr-4-en-3-one) is described, with the esterifying acids bearing an acetylenic or olefinic function in a chain of eight or nine carbon atoms, for evaluation as long-acting contraceptive agents.