Introduction of lysine residues on the light chain constant domain improves the labelling properties of a recombinant Fab fragment

Europium chelates provide a non-radioactive alternative forsensitive labelling of antibodies for diagnostic immunoassays.Lysine residues at antibody surfaces are ready targets for labellingby an isothiocyanate derivative of the europium chelate (Eu³⁺).Here the labelling efficiency of a recombinant anti-human -fetoprotein(hAFP) Fab fragment has been improved by increasing its lysinecontent by protein engineering. Molecular modelling was usedto identify three light chain constant domain surface arginineresidues, R154, R187 and R210, which were mutated to lysineresidues. The mutations did not influence the affinity of thelysine-enriched Fab fragment and its labelling efficiency wasfound to be 40% higher than that of the wildtype Fab fragmentWith low degree of labelling, the affinities of the two Fabfragments were identical and comparable with that of the originalmonoclonal anti-hAFP IgG. With a higher degree of labellingthe affinities of both Fab fragments decreased more than thatof the intact IgG since more lysine residues are available forlabelling in the additional heavy chain constant domains ofthe larger molecule. Electrostatic adsorption and covalent immobilizationof the Fab fragments were characterized by BIAcore^TM and thelysine-enriched Fab fragment was found to be more efficientlyimmobilized to an activated carboxymethyl surface.     

The prospects of the development of the method for monitoring of occupational exposure to some alkylating agents

DNA-adduct formation, depurination and imidazole ringopening were followed in vitro using styrene oxide, ethyleneimine and dimethyl sulfate. Depurination was found to be about 50 times faster in nucleosides than in double-stranded DNA. The half-lives of depurination in DNA were 3 times faster for 7-(2-aminoethyl)guanine as compared to 7-methyl- and 7-(2-hydroxy-2-phenylethyl)deoxyguanosine. In neutrally 7-methylguanine was released some 60 times faster than that of guanine and adenine. This apparent discrepancy in depurination between alkylated and intact bases suggests the possibility of developing a sensitive method for monitoring of DNA alkylations formed by electrophillic chemicals, which might be based on labelling of apurinic sites and utilized for in vivo studies as well.     

Suicides after pregnancy in Finland, 1987-94: register linkage study

OBJECTIVE: To determine rates of suicide associated with pregnancy by the type of pregnancy. DESIGN: Register linkage study. Information on suicides in women of reproductive age was linked with the Finnish birth, abortion, and hospital discharge registers to find out how many women who committed suicide had had a completed pregnancy during her last year of life. SETTING: Nationwide data from Finland. SUBJECTS: Women who committed suicide in 1987-94. RESULTS: There were 73 suicides associated with pregnancy, representing 5.4% of all suicides in women in this age group. The mean annual suicide rate was 11.3 per 100,000. The suicide rate associated with birth was significantly lower (5.9) and the rates associated with miscarriage (18.1) and induced abortion (34.7) were significantly higher than in the population. The risk associated with birth was higher among teenagers and that associated with abortion was increased in all age groups. Women who had committed a suicide tended to come from lower social classes and were more likely to be unmarried than other women who had had a completed pregnancy. CONCLUSIONS: The increased risk of suicide after an induced abortion indicates either common risk factors for both or harmful effects of induced abortion on mental health.     

A review of postmenopausal hormone therapy recommendations: potential for selection bias

OBJECTIVE: To survey the advice given to physicians in regard to postmenopausal hormone therapy, and thus to find out indirectly whether selective prescribing may have contributed to the findings of health effects. DATA SOURCES: Surveys on the health benefits of postmenopausal therapy were identified from previous reviews. Experts in various fields were consulted regarding relevant textbooks, and recommended booklists were used. METHODS OF STUDY SELECTION: Most of the surveys were conducted in the United States and concerned only estrogens; therefore, we focused on these data. Advice given to United States physicians on estrogen prescribing was abstracted from the Physicians' Desk Reference, five textbooks, and other sources written between 1945-1990. DATA EXTRACTION AND SYNTHESIS: In advice given to physicians, there was concern that estrogens could cause cancer. In addition, it was not recommended to give estrogens to women in poor general health. Advice concerning cardiovascular diseases and estrogens was contradictory and variable. CONCLUSIONS: If the advice observed reflects the prescribing practices of physicians, then surveys on the health impact of estrogen therapy may have underestimated the risk of breast cancer and overestimated the prevention of fractures. The impact on studies of cardiovascular diseases is less clear, and other mechanisms of selection bias may have been more important.     

Chromatin proteins of large and small brain nuclei

A simple fluorometric assay for monoamine oxidase (MAO) [EC 1.4.3.4] activity towards beta-phenylethylamine (PEA) was devised. The procedure consists in measuring the disappearance of PEA fluorometrically. The disappearance of PEA was completely inhibited by pargyline, a potent inhibitor of MAO. MAO activity for PEA was linear with 10 mg to 100 mg of liver tissue in 3 ml of reaction mixture for up to 90 min of incubation. Using this method, the V max values and the apparent Km values of MAO for PEA in several rat tissues were determined, and compared with those for benzylamine and 5-hydroxytryptamine (5-HT).     

On-line packing with boxes of different sizes

The packing of boxes distributed to different customers from a central depot is under consideration in this study. The boxes are of various sizes and they arrive on a conveyor belt one at a time. The on-line situation is assumed, which means that at each stage we only know the layout of the previous boxes on the partially filled pallet and the size of the box to be placed next but we have no information about the forthcoming ones. Further it is not allowed to move boxes already placed. The objective is to produce efficient and stable loads by an on-line packing algorithm, which can be implemented in automated systems. The algorithms we develop are based on the well-known first-fit and best-fit principles and their efficiency has been tested empirically.     

Activated Hepatic Stellate Cells in Hepatocellular Carcinoma: Their Role as a Potential Target for Future Therapies

Hepatocellular carcinoma (HCC) is a global healthcare challenge, which affects more than 815,000 new cases every year. Activated hepatic stellate cells (aHSCs) remain the principal cells that drive HCC onset and growth. aHSCs suppress the anti-tumor immune response through interaction with different immune cells. They also increase the deposition of the extracellular matrix proteins, challenging the reversion of fibrosis and increasing HCC growth and metastasis. Therapy for HCC was reported to activate HSCs, which could explain the low efficacy of current treatments. Conversely, recent studies aimed at the deactivation of HSCs show that they have been able to inhibit HCC growth. In this review article, we discuss the role of aHSCs in HCC pathophysiology and therapy. Finally, we provide suggestions for the experimental implementation of HSCs in HCC therapies.     

Self-medication of dyspepsia: how appropriate is it?

BACKGROUND: The scope of self-medication is increasing in many countries, and drugs for dyspepsia are a popular group for deregulative activities. This study investigated what kind of upper gastrointestinal symptoms people self-medicate and how appropriate this self-medication is. METHODS: An anonymous questionnaire was give to 50 consecutive customers buying antacids, alginates, or sucralfates in 10 pharmacies in the capital area in Finland in 1995. In half of the pharmacies the questionnaire was returned by mail, and in the other half the questionnaire was filled out at the pharmacy. The response rate was 53% (n = 292). RESULTS: The commonest reasons for self-medication were heartburn (88%), gastrointestinal pain (31%), and acid regurgitation (32%). Seventy-five per cent of respondents had used dyspepsia drugs for more than a year. The commonest way to self-medicate was to interchange regular and occasional use. Knowledge about the proper use of dyspepsia drugs was poor, and 6% of respondents had symptoms contraindicating self-medication but had not visited a physician during the past year. Patients more than 60 years old were especially at risk of potential inappropriate use. CONCLUSIONS: Over-the-counter (OTC) drugs for dyspepsia are likely to be used improperly. A physician's advice on the use of OTC dyspepsia drugs, in addition to detailed printed information about drug action and proper administration, would be important means to guarantee appropriate use of these drugs.     

Whole-Exome Sequencing Identifies a Novel Germline Variant in PTK7 Gene in Familial Colorectal Cancer

Colorectal cancer (CRC) is the third most frequently diagnosed malignancy worldwide. Only 5% of all CRC cases are due to germline mutations in known predisposition genes, and the remaining genetic burden still has to be discovered. In this study, we performed whole-exome sequencing on six members of a Polish family diagnosed with CRC and identified a novel germline variant in the protein tyrosine kinase 7 (inactive) gene (PTK7, ENST00000230419, V354M). Targeted screening of the variant in 1705 familial CRC cases and 1674 healthy elderly individuals identified the variant in an additional familial CRC case. Introduction of this variant in HT-29 cells resulted in increased cell proliferation, migration, and invasion; it also caused down-regulation of CREB, p21 and p53 mRNA and protein levels, and increased AKT phosphorylation. These changes indicated inhibition of apoptosis pathways and activation of AKT signaling. Our study confirmed the oncogenic function of PTK7 and supported its role in genetic predisposition of familial CRC.     

RNA metabolism in chick oviduct during oestrogen stimulation

The amount and the labelling of RNA were studied in chick oviduct after secondary stimulation of pre-treated chicks with oestrogen. The weight of the oviduct increases over 2 fold in 20 h of oestrogen action using a dose of 1 mg. During the hormonal treatment the amount of cytoplasmic and nuclear RNA increases about 3 and 1.5 fold, respectively. The amount of poly(A) containing RNA increases somewhat less than that of RNA lacking poly(A). The proportion of poly(A) containing species of the total cytoplasmic and nuclear RNA decreases from 5.8% to 4.4% and from 7.8% to 6.3%, respectively. Hydroxyurea largely prevents the effect of oestrogen in increasing the amount of RNA. Incorporation of [3H]uridine in RNA in vitro is stimulated about 2.5 fold in cytoplasmic RNA lacking poly(A) and only 1.2 fold in RNA containing poly(A) in 20 h of oestrogen action. In nuclear RNA the labelling of the two species is stimulated about 1.7 fold. Hydroxyurea fails to interfere with the oestrogen-induced stimulation in the labelling of RNA. Cytoplasmic and nuclear RNA containing poly(A) is distributed in a peak of about 20 S in agarose-acrylamide gels.