No abstract

A two-dimensional electrophoretic technique combining blue native polyacrylamide gel electrophoresis (BN-PAGE) with Tricine sodium dodecyl sulfate (SDS)-PAGE was previously used for the localization of oxidative phosphorylation (OXPHOS) defects in human diseases starting from biopsy or autopsy tissues (Schägger, H., Electrophoresis 1995, 16, 763-770). In the present work the technique was extended for the resolution of OXPHOS enzymes from platelets and tissue-cultured cells. Silver staining is required to detect the protein subunits of OXPHOS complexes in two-dimensional gels. However, the use of cultured cells has major implications for patients with mitochondrial encephalomyopathies since it will reduce the number of invasive muscle biopsies. The ease of isolating the platelet membrane glycoprotein complex from a few milliliters of blood makes it possible to analyze this complex and its protein subunits in bleeding disorders like Glanzmann's thrombasthenia.

Summary Starting with the isolation of thymine dimer derived from DNA of u.v.‐irradiated bacteria, a systematic investigation was begun from which it was concluded that the basis for the action of u.v.‐light is a photochemical dimerization. Depending on the dose of u.v.‐light and the conditions for hydrolysis of DNA and isolation procedures, other photoproducts of thymine were isolated. Experimental data for the conversion of cytosine to uracil are presented. By use of monochromatic u.v.‐light we could demonstrate that only intrastrand dimerization occurs. The radiation chemical behavior of uracil and cytosine was investigated with tritium labeled compounds. The mechanism for the various kinds of sensitization with 5‐halogen‐uracils, with dyes (thiopyronin, methylene blue, and pyronin) with carcinogenic hydrocarbons and with furocoumarins, which parallels partly the photochemical degradation of guanine is discussed. U.V.‐inactivated bacteria and polyuridylic acid are reactivated better by irradiation with visible light when hydrogen peroxide or uranyl acetate are present. The mechanism of this reaction is discussed. The absence of the α, β‐unsaturated double bond could be expected to protect the thymine molecule from dimerization; after incorporation of azathymine into the DNA, bacteria do indeed exhibit u.v.‐resistance. Moreover, after treatment of bacteria with hydrogen peroxide and visible light, by which the 5,6 double bond in the thymine and cytosine molecules is also attacked, bacteria become u.v.‐resistant. The specific alteration of uracil and cytosine by u.v.‐irradiation and of guanine through photodynamic action can be used as a simple method for sequence analysis of protein codons. It is also of use in determining the interaction between messenger‐RNA, transfer‐RNA and ribosomes during protein synthesis.

Four children with juvenile laryngeal papillomatosis were treated with human leukocyte interferon (IFN-alpha) by intravenous infusions. In three cases the clinical course of the disease was effected favorably. Serum IFN titers and (2'-5')oligo(A) synthetase (OAS) levels in lymphocytes of the patients were measured once a week during therapy. Levels of serum IFN determined 15 min after the end of 1 h infusions corresponded to only 15%-40% of the amount infused. Comparable OAS activities in the four children were measured before infusion, i.e., one, two, or three days, respectively, after the preceding infusion, though mean IFN titers of the patients differed from each other (200-400 and 400-600 u/ml). This suggests a saturation of the lymphocytes' antiviral system by these IFN levels. A discontinuation for two weeks in the early phase of IFN treatment, accompanied by a decrease of the OAS activity in the lymphocytes to the basal level, resulted in a deterioration of the patient's condition. A change in treatment schedule causing a decrease of OAS activity to a lower, though still elevated level, for six weeks until the present did not influence the course of therapy. Therefore, we suppose that the maintenance of elevated levels of OAS activity in lymphocytes for some months may be a necessary, even though not always a sufficient, criterion for a successful therapy schedule in IFN treatment of juvenile laryngeal papillomatosis. In addition, our results suggest OAS activity to be suitable in monitoring effects of an IFN therapy.