Oestrogen replacement therapy after hysterectomy.

OBJECTIVE--To identify those women who might benefit from oestrogen replacement after hysterectomy. DESIGN--Targeted health screening. SETTING--Large group practice. SUBJECTS--All women aged under 50 who had had a hysterectomy. MAIN OUTCOME MEASURES--Concentration of follicle stimulating hormone, symptom profile, uptake of oestrogen replacement therapy. RESULTS--145 of 1953 women aged 32-49 had had a hysterectomy. 35 of the 41 with bilateral oophorectomy and 27 of 104 with one or more ovaries conserved were taking oestrogen replacement. 62 of the 68 who had ovaries conserved and were not taking oestrogen replacement attended for review, of whom 14 had a follicle stimulating hormone concentration > or = 20 IU/l. 16 of the 19 women identified as potentially able to benefit from oestrogen replacement started treatment and were still on treatment at six months of follow up. CONCLUSION--Systematic review of women who had had a hysterectomy identified an important group who would potentially benefit from oestrogen replacement therapy.     

Plasmin desensitization of the PAR1 thrombin receptor: kinetics, sites of truncation, and implications for thrombolytic therapy

It has been hypothesized that protease-activated receptors may be activated and attenuated by more than one protease. Here, we explore a desensitization mechanism of the PAR1 thrombin receptor by anticoagulant proteases and provide an explanation to the enigma of why plasmin/tissue plasminogen activator (t-PA) can both activate and deactivate platelets prior to thrombin treatment. By using a soluble N-terminal exodomain (TR78) as a model for the full-length receptor, we were able to unambiguously compare cleavage rates and specificities among the serum proteases. Thrombin cleaves TR78 at the R41-S42 peptide bond with a kcat of 120 s-1 and a KM of 16 microM to produce TR62 (residues 42-103). We found that, of the anticoagulant proteases, only plasmin can rapidly truncate the soluble exodomain at the R70/K76/K82 sites located on a linker region that tethers the ligand to the body of the receptor. Plasmin cleavage of the TR78 exodomain is nearly equivalent to that of thrombin cleavage at R41 with similar rates (kcat = 30 s-1) and affinity (KM = 18 microM). Specificity was demonstrated since there is no observed cleavage at the five other potential plasmin-cleavage sites. Plasmin also cleaves the TR78 exodomain at the R41 thrombin-cleavage site generating transiently activated exodomain. We directly demonstrated that plasmin cleaves these same sites in full-length membrane-embedded receptor expressed in yeast and COS7 fibroblasts. The rate of plasmin truncation is similar between the extensively glycosylated COS7-expressed receptor and the nonglycosylated yeast-produced receptor. Mutation of the R70/K76/K82 sites to A70/A76/A82 eliminates plasmin truncation and desensitization of thrombin-dependent Ca2+ signaling and converts PAR1 into a plasmin-activated receptor with full agonist activity for plasmin. Plasmin does not desensitize the Ca2+ response of platelets or COS7 cells to SFLLRN consistent with intermolecular ligand-binding sites being located to the C-terminal side of K82. Truncation of the wild-type receptor at the C-terminal plasmin-cleavage sites removes the N-terminal tethered ligand or preligand, thereby providing an effective pathway for PAR1 desensitization in vivo.     

Use of frozen bagged seed inoculum for secondary metabolite and bioconversion processes at the pilot scale

Frozen bagged seed inoculum was prepared, thawed and tested for seven cultures. Thawing techniques were developed and other key influences on thawing rate were quantified; seed bag thawing without a water bath rarely required more than 4 to 5 h and was as short as 0.5 to 1 h for lower fill volume bags. Testing included growth of bagged seed as a function of bag fill volume (0.5, 1.0, 2.0, and 3.5 L), comparison of culture age at time of bagging, growth of bagged versus laboratory-prepared seed, productivity of production cultures derived from bagged versus laboratory-prepared seed, growth of bagged seed as a function of volume percent glycerol added at time of bagging, and growth of bagged seed as a function of frozen storage time and temperature. For each culture tested, conditions were developed such that seed tanks inoculated with bagged seed showed only minimal delay in attaining the target oxygen uptake rate (OUR) relative to seed tanks inoculated with laboratory-prepared inoculum. Although the bag fill volume did influence culture growth in some cases, bag fill volumes required were reasonable (typically 2.0 to 3.5 L) compared with laboratory seed inoculum volumes of 2.0 L. In the most remarkable example, frozen bagged seed was prepared from a second-stage seed-tank cultivation of Glarca lozoyensis, then thawed and inoculated into first-stage seed medium. It grew to the desired OUR in a similar timeframe as laboratory-prepared inoculum inoculated into first-stage seed medium. Thus, the frozen bagged seed replaced an existing laboratory inoculum preparation period of 7 days without an appreciable delay in either of the two subsequent seed-tank growth stages. Furthermore, productivities were found to be comparable for bagged-seed-derived and laboratory-seed-derived production cultivations for four different fermentation processes.     

Studies of acidosis in the ischaemic heart by phosphorus nuclear magnetic resonance.

1. Phosphorus-nuclear-magnetic-resonance measurements were made on perfused rat hearts at 37 degrees C. 2. With the improved sensitivity obtained by using a wide-bore 4.3 T superconducting magnet, spectra could be recorded in 1 min. 3. The concentrations of ATP, phosphocreatine and Pi and, from the position of the Pi resonance, the intracellular pH (pHi) were measured under a variety of conditions. 4. In a normal perfused heart pHi = 7.05 +/- 0.02 (mean +/- S.E.M. for seven hearts). 5. During global ischaemia pHi drops to 6.2 +/- 0.06 (mean +/- S.E.M.) in 13 min in a pseudoexponential decay with a rate constant of 0.25 min-1. 6. The relation between glycogen content and acidosis in ischaemia is studied in glycogen-depleted hearts. 7. Perfusion of hearts with a buffer containing 100 mM-Hepes before ischaemia gives a significant protective effect on the ischaemic myocardium. Intracellular pH and ATP and phosphocreatine concentrations decline more slowly under these conditions and metabolic recovery is observed on reperfusion after 30min of ischaemia at 37 degrees C. 8. The relation between acidosis and the export of protons is discussed and the significance of glycogenolysis in ischaemic acid production is evaluated.     

STUDY ON THE PROLIFERATIVE STATE OF HAEMOPOIETIC STEM CELLS (CFU)

Hydroxyurea was used to study the proliferation rate of haemopoietic stem cells (CFUJ in normal mice, after irradiation or transplantation into irradiated recipients. It was demonstrated that the proliferation rate of endogenous CFU_S (endo-CFU,) and exogenous CFU_S (exo-CFU_s) are identical. After irradiation (650 R) the surviving endo-CFU_s begin to proliferate immediately. By contrast exo-CFU, transplanted into the irradiated recipient mouse (850 R), begin to proliferate only after about 30 hr. However, injection of isoproterenol (which stimulates adenyl cyclase) or dibutyryl cyclic adenosine 3′,5′-monophosphate shortly after marrow cell graft, triggers the transplanted CFU_S into cell cycle as shown by an almost immediately increased sensitivity to hydroxyurea. Isoproterenol is capable of inducing DNA synthesis also in stem cells of normal mice but it takes about 20 hr before CFU, become to be increasingly sensitive to hydroxyurea.     

Distribution of phytochrome and chlorophyll-a/b-binding-protein mRNAs in etiolated Avena seedlings

In 4-d-old dark-grown oat (Avena sativa L.) seedlings, the majority of the type-I-phytochrome (phyA) mRNA was found within 10 mm of the tip of the coleoptile sheath and in the mesocotyl node; almost none was detected in the enclosed primary leaf. In contrast, chlorophyll-a/b-binding-protein (cab) mRNAs were found almost exclusively in the enclosed primary leaf and were barely detectable in total-RNA samples from the coleoptile sheath or mesocotyl node of red-light-treated etiolated seedlings. Separated, dark-grown primary leaves responded to a red-light treatment by increasing cab-mRNA abundance in the absence of the coleoptile sheath or mesocotyl node tissues.Abbreviations cab gene for chlorophyll-a/b-binding protein - kb kilobase - phyA gene for type-I-phytochrome protein We are grateful to the members of the laboratory Dave Higgs, Theresa Tirimanne, Dr. Dennis Byrne, Bruce Held, Linda Barnes, Dr. Isaac John, and Iffat Rahim, for their helpful discussions and critical review. This work was supported by USDA grant No. 88-37261-4196 and No. 91-37304-6397, the Iowa State University Biotechnology Program, and the Molecular, Cellular, and Developmental Biology Program.     

Immune deficiency in the X-linked lymphoproliferative syndrome. II. Immunoregulatory T cell defects

Surface phenotypic markers and the function of lymphocytes in patients affected with the X-linked lymphoproliferative syndrome (XLP) were studied. This syndrome is characterized by a defective response to infection with Epstein Barr virus (EBV). Normal numbers of B and T cells were detected with anti-Ig and monoclonal OKT3 antisera, respectively. T cell subset values, however, were persistently altered: cells reacting with OKT8 were significantly elevated in five of nine patients, accompanied by a slight decrease in the percentage of OKT4-positive cells, leading to abnormally low OKT4 to OKT8 ratios. One patient had a high OKT4 to OKT8 ratio due to low number of OKT8-positive cells. Lymphocytes from patients showed normal proliferation after stimulation with T and B cell mitogens. In contrast, Ig synthesis by lymphocytes after stimulation with B cell mitogens was markedly deficient: low or undetectable levels of one or all classes of Ig were detected, whereas cell lines established from EBV-infected B lymphocytes from patients produced normal quantities of Ig. These studies imply immune regulatory impairments in the patient with XLP.