3-chloro-1, 3, 5-pregnatriene derivatives with glucocorticoid activity

Novel synthetic glucocorticoid analogues were tested for receptor binding and glucocorticoid activity. They were of unusual structure, insofar as they had a 3-chloro rather than a 3-oxo function. 3-Chloro analogues of fluorinated glucocorticoids formed extremely stable complexes with the rat liver glucocorticoid receptor. 3-Chloro derivative of fluocinolone acetonide also had $\text{in vivo}$ glucocorticoid activity. It induced tyrosine aminotransferase in the liver and repressed thymidine kinase in the thymus very effectively. It is concluded that 3-chloro analogues may retain glucocorticoid activity as well as the ability to bind to the glucocorticoid receptor protein.     

The reaction of bovine pancreatic ribonuclease A with 6-chloropurineriboside 5'-monophosphate. Evidence on the existence of a phosphate-binding sub-site

The chemical modification of bovine pancreatic ribonuclease A by 6-chloropurine 9-beta-D-ribofuranosyl 5'-monophosphate was studied under several reaction conditions. The reaction, at pH 7.3, 40 degrees C and a nucleotide: enzyme molar ratio of 60, showed a high degree of specificity in comparison to those corresponding to the base or the nucleoside. The main derivative was isolated by means of CM-cellulose chromatography. Subtilisin cleavage of this derivative showed that the substitution had taken place in the S-peptide moiety. Tryptic digestion of the S-peptide indicated that a lysine residue had been modified. Enzymatic and physico-chemical considerations showed that the actual site of reaction was the alpha-amino group of Lys-1. The structural and kinetic properties of the derivative are consistent with the existence of a phosphate-binding sub-site near the N-terminal region of the enzyme.     

Quantification and distribution of troponin-T in cultures of chick embryo myocardiocytes.

We analyzed the distribution and labeling patterns of troponin-T, a protein involved in the regulation of striated muscle contraction, in myocardiocytes obtained from chick embryos in Hamburger and Hamilton's stage 25 and 39, and cultured for 8 days. Troponin-T expression was examined with indirect immunofluorescence, densitometry, fluorescence-activated cell sorting, electrophoresis and immunoblotting. The patterns of expression of troponin-T were compared with those of actin to determine possible correlations in different stages of chick embryo development and culture. Our findings show that in both stages of embryonic development, the cellular accumulation of troponin-T changed after 8 days of culture. Our results revealed a quantitative modification with time: after 4 days of culture there was a significant increase in this protein, followed by a slight additional increase after 8 days of culture. Flow cytometry findings confirmed these trends over time, showing a significant increase in positive cells after 4 days, followed by a smaller rise after 8 days of culture. In comparison with actin, this pattern was similar only in cells from Hamburger and Hamilton's stage 25 embryos.     

Propionyl-CoA synthetase in mammary gland and liver of cows

Propionyl-CoA synthetase of liver and mammary gland from calf and midlactation cow was investigated. No activity of this enzyme was detected in calf mammary gland, but it was detected in calf liver. Propionyl-CoA synthetase was found in both, liver and mammary gland of the cow, although mammary gland activity was about 25% of that found in liver. The effects of pH and temperature on enzyme activity and stability were also investigated in crude extracts of liver and mammary gland tissues. The results suggest a different behaviour of the enzyme from both origins. Kinetic studies of the enzyme were also carried out, showing differences, depending on the organ, in the apparent substrate KM values.     

Temperature and starvation affect the hemolymph acid-base balance of the xeric yellow scorpion,Leiurus quinquestriatus

Summary Water conservation is crucial for terrestrial animals such as scorpions which generally live in xeric habitats. One route of water loss is evaporation from their book lungs. In order to save water, scorpions may have a high resistance to gas exchange with the environment. If this is so then the partial pressure of CO₂ in their hemolymph, PCO₂, must be high. Does this affect their acid-base balance? Hemolymph PCO₂ and pH in normally-fed or starved desert-dwelling yellow scorpions Leiurus quinquestriatus were studied in vivo as functions of temperature. An ambient temperature increase (lasting at least 3 days) resulted in a rise of PCO₂ and a fall of pH, with thermal coefficients of 1.6 Torr · °C^-1 and-0.016 pH unit · °C^-1, respectively. The thermal coefficients for cell-free hemolymph studied in vitro were the same. At 28 °C, 3–6 weeks of starvation led to a 4.8 Torr increase in PCO₂ and a 0.056 unit decrease in pH. The in vivo PCO₂ values are among the highest, and pH values are the lowest of the terrestrial arthropods studied so far, e.g., at 28 °C they are 29 Torr and 7.15 pH respectively. It is argued that this particular acid-base balance with a marked hypercapnia is typical of a successful xeric air-breathing animal.Abbreviations ABB acid-base balance - PCO₂ partial pressure of CO₂ - PO₂ partial pressure of O₂     

Protein components of the erythromycin binding site in bacterial ribosomes

Two derivatives of erythromycin have been prepared carrying either an aryl azide or a 4-nitroguaiacol as a photoreactive group. Both derivatives bind to the specific erythromycin ribosomal site as shown by saturation and competition studies. The derivatives were isotopically labeled either with tritium or with 125I, and radioactivity is covalently incorporated to the ribosome upon irradiation at the appropriate wavelength. The ribosomal proteins labeled were identified by either mono- and two-dimensional gel electrophoresis or high performance liquid chromatography. It has been found that protein L22 is the protein mainly, and under some conditions exclusively, labeled by the erythromycin derivatives. These results were confirmed using ribosomes from erythromycin-resistant mutants having a protein L22 with modified electrophoretical mobility. Protein L15 is also labeled in both cases, and the aromatic azide derivative labels to a lesser extent proteins L2 and L4. Competition experiments with erythromycin indicate that labeling in protein L22, and probably in L15, is specific, while the specificity of labeling in proteins L2 and L4 is questionable. These results indicate that the erythromycin derivatives label different ribosomal proteins than the spiramycin type of macrolides (Tejedor, F., and Ballesta, J.P.G. (1985) Biochemistry 24, 467) suggesting that the binding sites of both macrolide types are probably not identical.     

Effect of dietary protein and energy intake on embryonic survival and gene expression in the uterine endometrium of early pregnant gilts

Porcine embryonic loss during early gestation is a serious problem in swine production. Improving embryonic survival can be achieved by maternal manipulation. Protein and energy are two major components of the diet, which play decisive roles in embryonic survival. This study was performed to evaluate the effects of enhancing maternal protein or energy intake on embryonic survival during early gestation in gilts and to explore the underlying mechanism. From day (d) 0 to 30 of gestation, 40 gilts (Landrace × York) were randomly allocated to 5 diets according to daily intake of low (L, National Research Council (NRC) recommendation for gestation gilts), medium (M, 20% higher than NRC) or high (H, 40% higher than NRC) CP or metabolisable energy (ME) (L_CPL_ME, M_CPL_ME, H_CPL_ME, L_CPH_ME, H_CPH_ME). Gilts were sacrificed on d 30 of gestation, and number of foetuses and corpora lutea, embryonic survival rate, uterine weight, and total volume of allantoic fluid were recorded or calculated. Gene expression was determined by Quantitative Real-time PCR (qPCR), western blot or immunohistochemistry. Results showed that increasing protein or ME intake significantly increased embryonic survival rate. Compared with diet L_CPL_ME, plasma progesterone (P4) concentration in diet L_CPH_ME increased at d 14 and d 30 of gestation. Progesterone receptor (PGR) was found not to be expressed in the epithelia but was strongly expressed in the stroma of the endometrium. Increasing protein or ME intake did not alter PGR expression in the endometrium. There was also no change in the amount of P4, hepatocyte growth factor, and fibroblast growth factor-7 in the endometrium. The mRNA abundance of cationic amino acid transporter 1 in the endometrium in diet L_CPH_ME and H_CPH_ME was significantly lower than in diet L_CPL_ME. Diet H_CPL_ME showed a tendency to increase neutral amino acid transporter 1 mRNA expression in the endometrium compared to diet L_CPL_ME (P = 0.087). In conclusion, increasing maternal protein or ME intake had a positive effect on the embryonic survival. Increased protein intake by 20 or 40% did not alter plasma P4 level, but increasing ME intake by 40% improved plasma P4 concentration at d 14 and 30 of gestation. Increasing maternal protein or ME intake did not induce PGR expression in the endometrium. Maternal protein and energy intake likely mediate transportation of cationic and neutral amino acids from mother to foetus to affect embryonic survival and development.     

Treefall Gaps and Regeneration Composition in the Laurel Forest of Anaga (Tenerife): a Matter of Size?

The size of treefall gaps is an important determinant of regeneration composition in tropical and temperate forests. Preliminary studies in the laurel forest of Tenerife have shown that small gaps (<100 m²) were the most numerous. However, due to this small size, no significant differences were found between regeneration in gaps and regeneration below the canopy. Because infrequent large gaps (>100 m²) are present in the laurel forest, we analyzed the regeneration in these large uncommon gaps, considering their potentially important role in the dynamics of the system. Our main hypothesis is that large gaps are important disturbance to ensure the regeneration and stablishment of shade intolerant species. Only five gaps larger than 100 m² (ranging from 125–268 m²) were found in the study area. Data from a further 20 small gaps (<100 m²), analysed in a previous study, was also included. Control plots were examined close to the gaps in order to determine regeneration below the closed canopy. We did not find a significant difference between regeneration density in the gaps (<100 m²) and regeneration below the canopy in the control plots. Contrary to our expectations, regeneration was lower in the large gaps than under the canopy. The open canopy in the large gaps increases light intensity, and has a negative effect on the germination and growth of shade-tolerant tree species like Viburnum tinus (although non-statistically significant); however, the increase in light intensity is not sufficient to stimulate the germination of shade-intolerant tree species. The effects of treefall gaps in the dynamics of the laurel forest of Anaga should be not considered as significant in comparison to other factors such as human disturbances or infrequent disturbances (land slides or hurricanes).