Biomedical subjects
Publications and source records attributed to M M Müller.
The complementary DNA coding for a lymphocyte-specific transcription factor binding to the DNA 'octamer' sequence TNATTTGCAT has been cloned. The nucleotide sequence shows homology to the homoeobox domain. Expression of this cDNA in HeLa cells is sufficient for a strong transcriptional activation of B-cell-specific promoters.
Homoeotic genes in Drosophila melanogaster are active in spatially restricted metameric domains and control the morphogenesis of segment-specific features such as legs or wings within these domains. They exert their function, according to the 'selector gene' hypothesis, by regulating the expression of subordinate genes. Homoeotic genes also control their own expression and the expression of each other. The proteins encoded by these genes contain a domain, called a homoeodomain, that is strongly conserved, and that shows homologies to proteins that bind DNA and regulate transcription. Homoeoproteins have been shown to bind specific DNA sequences. We show here that the Drosophila homoeotic genes Ultrabithorax (Ubx) and Abdominal-B (Abd-B) code for proteins that are capable of activating transcription of reporter genes linked to specific cis-regulatory target sequences in transfected mammalian cells. Their activity, as well as their target specificity, is similar to that of a mammalian lymphoid-specific octamer transcription factor, OTF-2, which was recently found to contain a homoeodomain.
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A case of adenine phosphoribosyltransferase deficiency in a 4.5-yr-old boy is described. A pedigree of the family, enzyme activities and kinetic data of the enzyme in the propositus and the carriers of the defect are presented. The amount of enzyme in the patient was about 2% of that in healthy subjects and correlated well with the amount of immunoreactive protein. Our data indicate that the patient's enzyme is not affected in its catalytic properties, but is made in far reduced amounts.
The diagnostic validity of multivariate combinations of alpha 1-antitrypsin, alpha 2-macroglobulin, C-reactive protein, complement C3, complement C4, neopterin in serum, and neopterin in urine as markers for acute cardiac allograft rejection and for differential diagnosis of rejection and infections was investigated in the follow-up of 37 patients with heart transplants. Rejection was diagnosed by endomyocardial biopsy. Infections were classified as 'no infection', 'viral infection', and 'bacterial, fungal or mixed infections'. Although there are significant differences between the mean levels of analytes, multivariate discriminant analysis does not provide an adequate discrimination of rejection and infection states. In separate rejection diagnosis, multivariate combinations of analytes cannot replace endomyocardial biopsy. However, a multivariate combination of alpha 1-antitrypsin, alpha 2-macroglobulin, C-reactive protein, C3, C4 in serum, and neopterin in urine can be used as a screening procedure to reduce the number of endomyocardial biopsies.
The influence of extracorporeal circulation (ECC) on human platelet adenine nucleotides has been studied in 28 coronary bypass patients before, during and after operation. An oscillating pattern of transient increases and decreases in total platelet ATP was observed following the sternotomy until the end of the operation. A highly significant increase in platelet ATP (20% +/- 14 of the pre-anaesthesia values) occurred during the first 24 h after surgery. Total platelet ADP however, did not show this oscillation nor was there any significant release of ADP from the platelets during ECC. Following collagen activation, increased amounts of 'releaseable ATP' were found after protaminization (124% +/- 38 of pre-anaesthesia values) (p less than 0.05), although whole blood aggregability was slightly reduced (89% +/- 18 of pre-anaesthesia values). This study indicates that 1) the metabolic ATP pool of circulating platelets underwent rapid changes during open heart surgery; 2) the majority of platelets did not release inert ADP during ECC; 3) there may be a compensatory enhancement of platelet function by other blood cells, which could explain the discrepancy between our aggregatory results in whole blood and those reported in platelet rich plasma (PRP) aggregometry.
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We have developed a vector, OVEC ("oligonucleotide vector") to study DNA sequences involved in the regulation of transcription in mammalian cells. This vector is equally suitable for studying expression in vivo after transfection into cells, or for transcription studies in vitro with cell-free extracts. Putative cis-acting DNA segments from enhancers or promoters can be inserted at a position immediately upstream of the TATA box and coding sequence of the rabbit beta-globin gene. A regulatory DNA segment can be tested by itself or in conjunction with an enhancer located either in an adjacent upstream position, or downstream of the beta-globin gene. S1 nuclease mapping can be used to study transcription from circular and linear templates and run-off transcription in vitro is also feasible. Transcripts from a reference globin gene with a small deletion around the transcription initiation site can be measured with the same S1 nuclease probe and thus serve as an internal standard. We demonstrate the usefulness of OVEC by inserting either short oligonucleotides comprising a metal responsive enhancer element, or the SV40 enhancer, directly upstream of the TATA box. Both constructs yield high levels of correctly initiated transcripts in a transient expression assay in HeLa cells. In a HeLa cell nuclear extract the SV40 enhancer stimulates transcription 40-fold.
We have detected a transcriptional enhancer sequence downstream from the adult beta-globin (beta A-globin) genes of chicken and duck. DNA segments from the beta-globin coding and flanking sequences were cloned into expression vectors containing the SV40 promoter linked to either the T antigen gene or the cat gene. The expression of these genes was measured in a chicken erythroid cell line transfected with the recombinant plasmids. We found that segments located about 400 bp downstream from the poly(A) site of both the chicken and duck beta A-globin genes (and about 1.5 kb upstream from the embryonic epsilon-globin gene) stimulate transcription of the test genes about five-fold. In chicken essentially the same segment was also found by others to act as an erythroid cell-specific enhancer [Hesse et al., Proc. Natl. Acad. Sci. USA 83 (1986) 4312-4316; Choi and Engel, Nature 323 (1986) 731-734]. The sequence containing this enhancer is conserved in evolution. A high degree of homology, reaching 84% in a segment 180 bp in length, was found between chicken and duck despite an evolutionary divergence of 70 myr.
Dopamine has already been recognized as an inhibiting factor of prolactin. On the other hand, the stimulators of prolactin are still judged diversely at the moment. In addition to hypothalamic TRH, serotonine as well as endogenous opiates and VIP are under discussion. The object of our studies was testing the function of VIP in directing prolactin secretion on three different groups of patients: 1. hyperprolactinaemic women of WHO-group VI; 2. patients who had undergone a TRH-test along with an endocrine check-up; 3. post partum in childbed. The determination was made from analyzing peripheral blood by radioimmunoassay. In 20 amenorrhoeic patients of WHO-group VI, their prolactin and VIP levels were determined between 8 and 10 a. m. Only 4 patients showed a VIP-concentration below 20 pg/ml (which represents the limit of detectability). The other 16 patients showed an increased concentration of VIP. In 15 cases there was an obvious correlation of VIP and prolactin concentrations according to the linear regression. In the course of a TRH-test to 20 and 40 minutes after initial blood examination prolactin and VIP were again determined. After 20 minutes all patients showed a threefold increase of the initial prolactin value. On the other hand, VIP increased in 3 cases only. Apart from all those patients who wanted to become pregnant, these three women--being postmenopausal--showed increased initial values already in the unstimulated condition. The third group of patients consisted of 26 women in childbed. Their prolactin and VIP were determined between the second and eight day post partum.(ABSTRACT TRUNCATED AT 250 WORDS)
Plasmatic coagulation parameters were studied in patients on parenteral cephalosporins with different hepatic pharmacokinetics. Sixty patients received either cefotaxime (4 g/day), ceftriaxone (2 g/day) or latamoxef (4 g/day) pre- and postoperatively for at least 5 days at random. They received parenteral nutrition without vitamin K supply and had no oral intake. A significant drop (p less than 0.05) in vitamin-K-dependent coagulation factors was recorded in patients treated with latamoxef, while patients receiving ceftriaxone and cefotaxime did not exhibit a significant change in their plasmatic coagulation parameters. Interference of some cephalosporins with the vitamin-K-dependent hepatic metabolism of clotting factors seems to be likely, rather than a suppression of intestinal vitamin K production by the intestinal microflora.
To evaluate the possibility of providing, in vitro, an endothelial lining for artificial hearts, we cultivated adult autologous endothelial cells on two polyurethane and two silicone rubber surfaces. Over the ensuing 11-day period, we investigated the resulting cell proliferation and morphology by means of scanning electron and light microscopy. On the silicone rubber surfaces, seeding of 200,000 human saphenous vein endothelial cells per cm(2) produced an ideal cobblestone monolayer within a single day. In contrast, the polyurethane surfaces displayed an uneven, patchy distribution of endothelial cells. Scanning electron microscopy revealed microvilli and marginal overlapping in both groups. After the first day, the cell count on the polyurethane surfaces increased, whereas the count on the silicone rubber surfaces decreased. Morphologic investigations revealed that the ideally shaped cells initially on the silicone rubber had begun to overspread and subsequently to become detached, leaving denuded spheroid areas. Moreover, cultivation for 11 days on the polyurethane surfaces resulted in an unevenness of cell distribution that far exceeded the unevenness seen on the first day. Thus, despite the fact that materials with a high surface tension (such as silicone rubbers) seem to be ideal for initial cell spreading, subsequent cultivation results in cell detachment and death. On materials with a lower surface tension (such as polyurethanes), the less differentiated monolayers do at least proliferate, although their morphology remains unsatisfactory. Even if adult human endothelial cells should prove shear-stress-resistant, a minimum of 6 to 8 weeks would be required to establish autologous endothelial cell monolayers on the inner surface. Therefore, the endothelialization of artificial hearts is not possible when such hearts are used for urgent "bridging" before cardiac transplantation.
Fifteen men undergoing extracorporeal circulation for aorta-coronary bypass grafting were investigated for alterations of the plasma levels of cross-linked fibrin degradation products, protein C, free protein S, coagulation factor II, immunoglobulin G, and albumin. Although all patients were given heparin, a progressive increase of cross-linked fibrin degradation products was recorded during extracorporeal circulation, which indicates an activation of the plasmatic coagulation system. This increase was most pronounced in the late phase of extracorporeal circulation after reperfusion of the lung and in the early postoperative period. The levels of all other investigated plasma proteins decreased drastically after the patient was connected to the bypass circuit, which was primed with saline solution. These levels increased after termination of extracorporeal circulation and administration of fresh-frozen plasma. To study the consumption of protein C, protein S, and factor II during extracorporeal circulation, we formed ratios of the values of these parameters to the value of immunoglobulin G. After this volume correction, protein C was found to decrease significantly in the late phase of extracorporeal circulation, remaining low in the early postoperative period; protein S increased significantly soon after the onset of extracorporeal circulation and decreased after termination of extracorporeal circulation; factor II was unaffected by extracorporeal circulation, showing only a slight, insignificant increase in the postoperative phase. These results suggest a disturbance of the protein C system by extracorporeal circulation, which is possibly linked to the reported high bleeding tendency in patients undergoing operations with extracorporeal circulation.
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Enzyme activities of purine catabolism and salvage, the concentrations of high-energy phosphates and the reutilisation of purine bases and purine nucleosides were studied in rat heart myoblasts and myocytes. Rat heart myoblasts H9c2(2-1) were grown in Dulbecco's modified Eagle's minimum essential medium supplemented with 10% fetal calf serum. Reduction of fetal calf serum to 2% for 1 week resulted in a differentiation into myocytes with respect to their morphological features and their enzyme pattern. In differentiated myocytes, activity of 5'-nucleotidase was increased more than 2-fold, and AMP deaminase and creatine kinase activities were more than 10-fold elevated. The concentration of creatine phosphate in differentiated myocytes was doubled compared to that in myoblasts. The uptake into myoblasts and myocytes and the incorporation into adenine nucleotides was highest using adenosine, inosine and adenine uptake rates were intermediate, and hypoxanthine was utilised least. Differentiation of myoblasts into myocytes resulted in a slightly lower overall uptake of adenosine and adenine, whereas about 40% more inosine and hypoxanthine were utilised by myocytes. Increasing the phosphate concentration in the incubation medium up to 50 mmol/l resulted in a stimulation of uptake of all purine compounds tested. This stimulation was more pronounced in myoblasts.
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