TGF-beta 1-induced inhibition of mouse mammary ductal growth: developmental specificity and characterization

TGF-beta 1, implanted into growing mouse mammary glands, was previously shown to inhibit ductal growth in an apparently normal and fully reversible manner. In this report we extend these findings to show that TGF-beta 1 inhibition is highly specific. In pregnant or hormone-treated mice, doses of TGF-beta 1 that were capable of fully inhibiting ductal elongation had little effect on the proliferation of lobuloalveolar structures. Additionally, the inhibitory action of TGF-beta 1 on ducts is epithelium-specific, resulting in cessation of DNA synthesis in the rapidly proliferating epithelium of mammary end buds, but does not inhibit DNA synthesis in the stroma surrounding the end buds. At the cellular level, transplant studies showed that TGF-beta 1 inhibited the regeneration of mammary ductal cells when implanted into mammary gland-free fat pads by suppressing the formation of new end buds, without inhibiting maintenance DNA synthesis in ductal lumenal epithelium; this observation indicates the potential of TGF-beta 1 to maintain patterning by suppressing adventitious lateral branching. The time-course of TGF-beta 1 inhibition of end buds was rapid, with cessation of DNA synthesis by 12 hr, followed by loss of the stem cell (cap cell) layer. The question of glandular exposure to TGF-beta 1 administered in EVAc implants was also investigated. Incorporation of TGF-beta 1 into EVAc was found not to degrade the hormone, while the release kinetics of the ligand from implants, its retention in the gland, and the demonstrable zone of exposure were consistent with observed inhibitory effects. These results support the hypothesis that TGF-beta 1 is a natural regulator of mammary ductal growth.     

Longitudinal age changes in a fishermen population of West Bengal

The longitudinal anthropological age changes in a sample of coastal fishermen in West Bengal are discussed. The results show that some traits follow the trends observed in ageing of Western populations, for instance nasal length and breadth continue to increase throughout life in both sexes. Stature follows also the trend generally observed: a decrease occurs indeed from 35 years of age in both sexes, the decline is becoming higher with increasing age. The typical ecological conditions of our population influence the rate of change in other traits, such as weight and biacromial diameter. Weight is known to be influenced by the nutritional status and the way of life. It explains that in our study of males an early increase of weight and biacromial diameter is observed first and a decrease afterwards. In females, however, a decrease is observed already from 35 years of age. This kind of analysis was only possible with the help of longitudinal data and in a well defined population: only longitudinal data is free from the effect of secular changes and of selective death.     

Evidence that ATP-dependent Ca2+ transport in rat parotid microsomal membranes requires charge compensation.

ATP-dependent Ca2+ transport was investigated in a rat parotid microsomal-membrane preparation enriched in endoplasmic reticulum. Ca2+ uptake, in KCl medium, was rapid, linear with time up to 20 s, and unaffected by the mitochondrial inhibitors NaN3 and oligomycin. This Ca2+ uptake followed Michaelis-Menten kinetics, and was of high affinity (Km approximately 38 nM) and high capacity (approximately 30 nmol/min per mg of protein). In the presence of oxalate, Ca2+ uptake continued to increase for at least 5 min, reaching an intravesicular accumulation approx. 10 times higher than without oxalate. Ca2+ uptake was dependent on univalent cations in the order K+ = Na+ greater than trimethylammonium+ greater than mannitol and univalent anions in the order Cl- greater than acetate- greater than Br- = gluconate- = NO3- greater than SCN-. Ca2+ uptake was not elevated if membranes were incubated in the presence of a lipophilic anion (NO3-) and carbonyl cyanide p-trifluoromethoxyphenylhydrazone. Ca2+ transport was altered by changes in the K+-diffusion potential of the membranes. A relatively negative K+-diffusion potential increased the initial rate of Ca2+ accumulation, whereas a relatively positive potential decreased Ca2+ accumulation. In the presence of an outwardly directed K+ gradient, nigericin had no effect on Ca2+ uptake. In aggregate, these studies suggest that the ATP-dependent Ca2+-transport mechanism present in rat parotid microsomal membranes exhibits an electrogenic Ca2+ flux which requires the movement of other ions for charge compensation.     

Induction and modification of dinitrogenase reductase in the unicellular cyanobacterium Synechocystis BO 8402

Oxygen is an important regulatory factor of nitrogenase induced in a unicellular cyanobacterium, Synechocystis BO 8402, during nitrogen starvation. Synthesis of the enzyme is limited by the efficiency of the cells to remove oxygen by respiration, supported by hydrogenases and, in the light, by inhibition of photosynthesis. With a polyclonal antibody against dinitrogenase reductase (the Fe protein of nitrogenase) a single polypeptide is detected, indicative of an active dimeric enzyme in dense cell suspensions. Inhibition of nitrogenase by addition of oxygen is accompanied by the appearance of a second polypeptide of the Fe protein having a 1.5 kDa higher molecular weight. This disappears upon removal of oxygen from the gas phase while nitrogenase activity is restored. No protein synthesis is required indicating that a fraction of the existing polypeptides is reversibly modified in response to oxygen. After induction of nitrogenase activity in dilute culture suspensions, both forms of the Fe-protein are found in variable amounts possibly due to oxygen contamination during the experiment.Abbreviations CAM chloramphenicol - Chl chlorophyll a - CHO carbohydrates - DCMU 3,4-dichlorophenyl-1,1-dimethylurea (diuron) - kDa kilodalton - SDS sodium dodecylsulphate     

Molecular genetic analysis of 67 patients with duchenne/becker muscular dystrophy

Summary A total of 56 Duchenne muscular dystrophy (DMD) patients and 11 Becker muscular dystrophy (BMD) patients was analyzed by extended multiplex amplification of the DMD/BMD gene; deletions were found in 60% of these patients. The data obtained were used to test the frameshift hypothesis and to compare the distribution of familial versus isolated cases. A significant correlation was found between deletions and isolated cases. Additional experiments were performed in order to determine the deletion breakpoints more precisely. These data are a prerequisite for carrier analysis in the respective families by detection or exclusion of aberrant cDNA fragments derived from ectopic lymphocyte RNA. This diagnostic technique is illustrated by 5 examples.     

Activation of the inositol trisphosphate second messenger system by cAMP in a mouse fibroblast cell line

Intracellular Ca²⁺ mobilization events were assessed in mouse L cells, which contain native prostaglandin E₁ receptors and transfected human ₂ adrenergic receptors. Both Fura2 (single cell measurements) and Quin 2, (cuvette assays) were used to determine [Ca²⁺]_i levels. Our results demonstrate that in the transfected cells there is a dose-dependent increase in [Ca²⁺]_i in response to isoproterenol (0.1 nM–100 nM), which is inhibited by the -adrenergic antagonist, propranolol, and is a result of intracellular Ca²⁺ release. [Ca²⁺]₁ in these cells was also increased by prostaglandin E₁, 8 bromo cyclic AMP, and aluminum fluoride. Both 8 bromo cAMP and isoproterenol induced a rapid increase in the levels of IP₁, IP₂, and IP₃. The data presented demonstrate that the elevation of intracellular cyclic AMP induces an increase in IP₃ production which leads to an elevation in [Ca²⁺];. We propose that this cyclic AMP dependent activation of the IP₃ generating system occurs at a post-receptor site.Abbreviations cAMP Adenosine Cyclic 3-5-Monophosphate - [Ca²⁺]_i intracellular [Ca²⁺]_i - 8 Br cAMP 8 Bromo Adenosine Cyclic 3-5-Monophosphate - DAG Diacylglycerol - EGTA] [Ethylene Bis (oxyethylenenitrilo)] Tetracetic acid - BSA Bovine Serum Albumin - HBSS-H Hanks' Balanced Salt Solution buffered with HEPES to pH 7.4 - HEPES 4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid - PIP₂ Phosphatidylinositol 4,5-bisphosphate - IP₂ Inositol 4 Phosphate - IP₂ Inositol 4,5 Bisphosphate - IP₃ Inositol Trisphosphate - PGE₁ Prostaglandin E₁ - PBS Phosphate Buffered Saline Solution