Interacting role of thyroxine and growth hormone in the hepatic synthesis of alpha 2u-globulin and its messenger RNA

Hypophysectomy completely abolishes and thyroidectomy results in a 90% reduction in the hepatic content of alpha 2u-globulin and its mRNA in the male rat. Thyroid hormone is also known to be required for the synthesis and secretion of pituitary growth hormone. In the hypothyroid rat either thyroxine or growth hormone was found to increase the activity and number of sequences of the mRNA for alpha 2u-globulin (measured by translational assay and hybridizational analysis with a cloned cDNA probe) to the euthyroid level. Treatment of hypophysectomized rats with a hormone combination containing growth hormone but not thyroxine increased the hepatic level of the mRNA for alpha 2u-globulin to that of normal animals. From these results we conclude that thyroxine indirectly influences the hepatic concentration of the mRNA for alpha 2u-globulin through its effect on pituitary growth hormone. Although administration of growth hormone to hypothyroid animals raised the hepatic concentration of alpha 2u-globulin mRNA to the euthyroid level, synthesis of alpha 2u-globulin remained low (50% of the normal). Complete recovery of alpha 2u-globulin synthesis required thyroxine. Therefore, in addition to an indirect effect on the hepatic level of alpha 2u-globulin mRNA, thyroxine also directly influences the synthesis of this protein. This direct effect of thyroxine on alpha 2u-globulin synthesis seems to be exerted at a step distal to the formation of mature mRNA.     

Characterisation of Trypanosoma cruzi populations by dna polymorphism of the cruzipain gene detected by single-stranded dna conformation polymorphism (SSCP) and direct sequencing

Fifty fresh isolates of Trypanosoma cruzi from Triatoma dimidiata vectors and 31 from patients with Chagas disease were analysed for DNA polymorphisms within the 432-bp core region of the cruzipain gene which encodes the active site of cathepsin L-like cystein proteinase. The cruzipain gene showed signs of polymorphism consisting of four different DNA sequences in Central and South American isolates of T. cruzi. The PCR fragments of Guatemalan isolates could be divided into three groups, Groups 1, 2 and 3, based on different patterns of single-stranded DNA conformation polymorphism. All of the strains isolated from Brazil, Chile, and Paraguay, except for the CL strain, showed a Group 4 pattern. Two to four isolates from each group were analysed by cloning and sequencing. A silent mutation occurred between Groups 1 and 2, and five nucleotides and two aa substitutions were detected between Groups 1 and 3. The DNA sequence of Group 4 contained five nucleotides and one aa substitution from Group 1. All of the DNA sequences corresponded well with the single-stranded DNA conformation polymorphism. The Group 1 isolates, the majority in the Guatemalan population (70/81, 86.4%), were isolated from both triatomines and humans, but Group 3 were isolated only from humans. Moreover, the Group 2 isolates were detected only in triatomine vectors (9/50; 18%), but never in humans (0/32, P<0.05) suggesting that this group has an independent life-cycle in sylvatic animals and is maintained by reservoir hosts other than humans.     

Luminescence immunoassay of pollen allergens on air sampling polytetrafluoroethylene filters.

A new method for quantification of airborne birch and grass pollen allergens collected on porous polytetrafluoroethylene filters has been developed. In this method, the allergens firmly adsorbed to the sampling filter of 25 mm in diameter are reacted with specific antibodies conjugated to alkaline phosphatase, generating a matrix-bound allergen-antibody-phosphatase complex. The filter is then floated on a chemiluminescent enzyme substrate solution. The light intensity of the product is linearly related to the amount of allergen over a large mass range, 0-1000 SQ (1 SQ is about 250 pg of protein). This direct on sampling filter in solution (DOSIS) technique demonstrated intra-assay precisions between 6-16% and 11-15% for the levels of 1-100 SQ units of grass allergen Phl p 5 and 4-400 SQ units of birch allergen Bet v 1, respectively. The limits of quantification for the corresponding allergens were estimated to 0.5 and 2 SQ units. Application of DOSIS to analysis of the grass pollen allergen concentrations of outdoor air for 12 days in July 1998 revealed a correlation coefficient of 0.69 between pollen grain and allergen concentrations for the dry weather period. After rainy days large amounts of grass allergens were present even in the absence of pollen grains.     

Elementary steps in protein folding.

The mechanism of protein folding is under intense theoretical and experimental investigation. From stopped-flow mixing experiments we have detailed knowledge of processes slower than about 1 ms, but until recently little was known about folding and unfolding reactions on the microsecond to nanosecond time scale. The use of novel techniques allowed to explore the elementary steps in protein folding, such as intrachain diffusion and formation of alpha-helices, beta-hairpins and loop structures. This brief review discusses the time scales of these early elementary events which are crucial for the understanding of how proteins fold.     

Direct microsensor measurement of nitric oxide production by the osteoclast.

Nitric oxide (NO) triggers marked osteoclast retraction which closely resembles that due to Ca2+. The effect of Ca2+ has been attributed to a stimulated release of NO. Here, we show for the first time, by direct measurement with a microsensor, that osteoclasts do indeed produce NO and that this production is enhanced by a high Ca2+. We also show that the Ca2+ ionophore, A23187, mimics the latter. Furthermore, osteoclasts on dentine produce more NO than osteoclasts on glass and NO release from dentine-plated osteoclasts is much less sensitive to stimulation by Ca2+. Finally, the microsomal Ca2+ store-depleting agent, thapsigargin, attenuates NO release only from osteoclasts on glass, suggesting that stored Ca2+ has the dominant effect in modulating NO release from non-resorbing cells. NO is a powerful inhibitor of bone resorption: a direct demonstration of its production is therefore strong evidence for a role in modulating osteoclast function.