Otto von Deimling

Die Darstellung phosphatfreisetzender Enzyme mittels Schwermetall-Simultan-Methoden

ZusammenfassungPhosphatfreisetzende Enzyme werden histochemisch nach einer Simultan-methode dargestellt, bei welcher Bleikomplexe verwendet werden. Die Vorteile der Verwendung von Bleikomplexen werden aufgeführt: Im alkalischen Bereich kann gegenüber den Gomori-Methoden ein Reaktionsschritt eingespart werden, im neutralen und sauren Bereich treten trotz relativ hoher Blei -und Organophosphatkonzentrationen keine Störungen durch unspezifische Niederschläge auf. Vorschriften zur histochemischen Darstellung von sechs phosphatfreisetzenden Enzymen werden als Beispiele aufgeführt und mit Abbildungen belegt.SummaryPhosphate liberating enzymes are demonstrated by a simultaneous heavy metal method whereby lead complexes are used. The advantages of using lead complexes are described: In the alkaline range one reaction stage can be left out in contrast to the Gomori method. In the neutral and acid range no disturbances occur through unspezific deposits despite relatively high concentrations of lead and organic phosphates. Instructions for the histochemical demonstration of six enzymes which liberate anorganic phosphate are given as examples and illustrated.

The recombinant congenic strains—a novel genetic tool applied to the study of colon tumor development in the mouse

The development of tumors in mice is under multigenic control, but, in spite of considerable efforts, the identification of the genes involved has so far been unsuccessful, because of the insufficient resolution power of the available genetic tools. Therefore, a novel genetic tool, the RC (Recombinant Congenic) strains system, was designed. In this system, a series of RC strains is produced from two inbred strains, a “background” strain and a “donor” strain. Each RC strain contains a different small subset of genes from the donor strain and the majority of genes from the background strain. As a consequence, the individual genes of the donor strain which are involved in the genetic control of a multigenic trait, become separated into different RC strains, where they can be identified and studied individually. One of the RC strains series which we produced is made from the parental strains BALB/cHeA (background strain) and STS/A (donor strain). We describe the genetic composition of this BALB/cHeA-C-STS/A (CcS/Dem) series and show, using 45 genetic autosomal markers, that it does not deviate from the theoretical expectation. We studied the usefulness of the CcS/Dem RC strains for analysis of the genetics of colon tumor development. The two parental strains, BALB/cHeA and STS/A, are relatively resistant and highly susceptible, respectively, to the induction of colon tumors by 1,2-dimethylhydrazine (DMH). The individual RC strains differ widely in colon tumor development after DMH treatment; some are highly susceptible, while others are very resistant. This indicates that a limited number of genes with a major effect are responsible for the high susceptibility of the STS strain. Consequently, these genes can be mapped by further analysis of the susceptible RC strains. The differences between the RC strains were not limited to the number of tumors, but the RC strains differed also in size of the tumors and the relative susceptibility of the two sexes. Our data indicate that the number of tumors and the size of tumors are not controlled by the same genes. The genetics of these different aspects of colon tumorigenesis can also be studied by the RC strains. The DMH-treated mice of the parental strains and the RC strains also developed anal tumors and haemangiomas in varying numbers. The strain distribution pattern (SDP) of susceptibility for each of the three types of tumors induced by DMH is different, indicating that development of these tumors is under control of different, largely non-overlapping, sets of genes. Thus, with a single series of RC strains, genes involved in tumorigenesis in various organs and tissues can be studied separately. These results indicate that the novel genetic tool, the RC strain system, offers new possibilities for analysis of the multigenic control of tumor development.

Die histochemische Lokalisation der Phosphatase in der Tubulusepithelzelle der Mäuseniere im elektronenmikroskopischen Bild

An der Niere der Albinomaus wurde nach Vorfixierung mit Osmiumtetroxyd eine modifizierte Gomori-Reaktion auf Phosphatasen durchgefuhrt. Als Fallungsmittel der bei dem Test freigesetzten Orthophosphorsaure wurde ein Bleisalz verwendet. Damit konnte im elektronenmikroskopischen Bild am Enzymort ein feinkorniger Niederschlag mit hohem Kontrast erzielt werden. Als Sitz der beiph 7,6–7,7 wirksamen Phosphatasen konnten die Membran des Burstensaums und die basalen Einstulpungen der Zellmembran der Tubulusepithelzellen erkannt werden. Enzymatisch aktiv wurden ferner die Zellmembranen der Kapillarendothelien des Tubulus und des Glomerulus sowie die Deckzellfortsatze (Microvilli) des Glomerulus befunden. Das Problem der intrazellularen Lokalisation von Phosphatasen wird diskutiert.ZusammenfassungAn der Niere der Albinomaus wurde nach Vorfixierung mit Osmiumtetroxyd eine modifizierte Gomori-Reaktion auf Phosphatasen durchgeführt. Als Fällungsmittel der bei dem Test freigesetzten Orthophosphorsäure wurde ein Bleisalz verwendet. Damit konnte im elektronenmikroskopischen Bild am Enzymort ein feinkörniger Niederschlag mit hohem Kontrast erzielt werden. Als Sitz der beiph 7,6–7,7 wirksamen Phosphatasen konnten die Membran des Bürstensaums und die basalen Einstülpungen der Zellmembran der Tubulusepithelzellen erkannt werden. Enzymatisch aktiv wurden ferner die Zellmembranen der Kapillarendothelien des Tubulus und des Glomerulus sowie die Deckzellfortsätze (Microvilli) des Glomerulus befunden.Das Problem der intrazellulären Lokalisation von Phosphatasen wird diskutiert.SummaryA modified Gomori-reaction on phosphatases of the albino-mouse kidney was performed. The method of prefixation with osmiumtetroxyde was used. The orthophosphoric acid, liberated in the test and cristallized by lead salt, yielded a fine granulated sediment with a high contrast at the location of enzyme in the electronmicroscopic picture. The phosphatases, active at aph of 7,6–7,7, could be located in the membranes of the brushborder and in the basal invaginations of the membranes of the tubuli cells. Furthermore an enzymatic activity was observed in the membranes of the capillary endothelium cells in the tubulus as well as in the glomerulus and in the microvilli of the glomerulus.Problems resulting from the test for the phosphatase were discussed.

Biochemical markers in inbred strains of the rat (Rattus norvegicus).

Klaus Bender 1, M a r k Adams 2, Peter R. Baverstock 2, Maria den Bieman 3, Siegbert Bissbort 1, Rad im Brdi~ka 4, Geoffrey W. Butcher 5, Dona ld V. Cramer 6, Otto yon Deimling 7, Michael F .W. Festing 8, Eberhard Gtinther 9, Rona ld D. G u t t m a n n 1°, Hans J. Hedrich 11, Philip B. Kendall 12, Reinhard Kluge i t , Ren6 Moutier 13, Babette Simon 7, James E. W o m a c k ~4, Junzo Yamada ~5, and Bert van Zutphen 3

Methodische Untersuchungen zur quantitativen und histologischen Verteilung der alkalischen Phosphatase in den Hauptstücken der Rattenniere

Summary1.A preparation method to get freeze-dried material of kidney cortex and kidney subcortex respectively on a 10 mg scale is described.2.The specific activity of alkaline phosphatase in this material was determined. The quantitative values were compared with the results of the histochemical demonstration of the phosphatase in the same kidneys.3.A satisfying agreement between the histochemical results and the quantitative data could be shown. The quantitative method is more sensible than the method of comparing histochemical preparations. Furthermore, the latter method is not strictly objective, since the histochemically demonstrated distribution of alkaline phosphatase considerably depends on the incubation conditions as pH and time of incubation.Zusammenfassung1.Für Rattennieren wurde eine Präparationsweise angegeben, mit welcher gefriergetrocknetes Material aus Rinde einerseits und Markaußenstreifen andererseits im 10 mg-Maßstab getrennt gewonnen werden kann.2.Die spezifische Aktivität der alkalischen Phosphatase in diesem Material wurde bestimmt. Die quantitativen Werte wurden anhand charakteristischer Beispiele den Resultaten der histochemischen Phosphatasedarstellung in den gleichen Nieren gegenübergestellt.3.Es wurde gezeigt, daß die histochemischen Resultate in einem befriedigenden Einklang mit den quantitativen Meßwerten zu bringen sind. Die quantitative Methode ist empfindlicher als das histochemische Vergleichsverfahren. Außerdem ist das histochemische Vergleichsverfahren nicht streng objektiv, da das Bild der Phosphataseverteilung im histochemischen Präparat in beträchtlichem Maß von den Inkubationsbedingungen wie pH-Wert und Inkubationsdauer abhängig ist.

Hormonabhngige Enzymverteilung in Geweben: I. Histochemische Untersuchungen ber die Geschlechtsunterschiede der alkalischen Nierenphosphatase bei erwachsenen Ratten

ZusammenfassungEs wurde gezeigt, daß sich männliche und weibliche Rattennieren durch die Verteilung und die Intensität ihrer alkalischen Phosphataseaktivität unterscheiden. Nach Kastration und nachfolgender östradiolbehandlung nimmt die Niere männlicher Ratten eine für weibliche Tiere typische Verteilung an.

Esterase. VIII. Histochemische, elektrophoretische und quantitative Untersuchungen zum Einfluß von Phenobarbital auf die Leberesterase der Maus

SummaryThe influence of phenobarbital upon the esterase activity of liver was studied in 48 mice of either sex by quantitative, electrophoretic and histochemical methods. 1.The esterase of mouse liver shows primary histochemical, quantitative and electrophoretic differences related to sex.2.A higher activity of esterase has been observed in both sexes under the influence of phenobarbital. Males show a slightly higher phenobarbital effect than females. The male type of banding pattern shifts towards the female type.3.It seems possible that the sex-depending and the phenobarbital effect are caused by the same mechanism.ZusammenfassungBei 48 Mäusen (NMRI) beiderlei Geschlechts wurde der Einfluß von Phenobarbital auf die Leberesterase histochemisch, quantitativ und disk-elektrophoretisch untersucht. 1.Die Leberesterase zeigt primäre histochemische, quantitative und elektrophoretische Geschlechtsunterschiede.2.Phenobarbital führt bei beiden Geschlechtern zu einer Erhöhung der Esteraseaktivität. Der Effekt ist bei männlichen Tieren etwas größer als bei weiblichen Tieren. Bei männlichen Tieren wird der Typ des Bandenmusters demjenigen weiblicher Tiere angeglichen.3.Es besteht der Verdacht, daß der Geschlechtseffekt durch den gleichen Wirkungsmechanismus erzeugt wird wie der Phenobarbitaleffekt.

Gene mapping on mouse chromosome 8 by interspecific crosses: New data on a linkage group conserved on human chromosome 16q

A large conserved linkage group exists on mouse chromosome 8 and human chromosome 16q, including the loci for chymotrypsinogen B (Ctrb), haptoglobin (Hp), lecithin:cholesterol acyltransferase (Lcat), metallothionein-1,-2 (Mt-1,-2), tyrosine aminotransferase (Tat), and uvomorulin (Um). Using cloned gene probes, these six loci were mapped in M. m. domesticus X M. spretus interspecific crosses relative to a number of chromosome 8 anchor loci resulting in the gene order Es-1,Es-9-Mt-1,-2-Got-2-Es-2,Es-7,Lcat,Um-Hp,Tat,Ctrb-e. These results complement earlier studies and redefine the conserved segment on mouse chromosome 8, previously defined by the Hp-Tat interval, by the 24-cM interval between Mt-1,-2 and the conserved locus for adenine phosphoribosyltransferase, Aprt, mapped at 25 cM from Es-1 by T. B. Nesterova, P. M. Borodin, S. M. Zakian, and O. L. Serov (1987, Biochem. Genet. 25: 563-568). Within this segment, the gene order appears the same in man and mouse. While map distances between HP-TAT,HP-CTRB, and TAT-CTRB of respectively 7, 11, and 9 cM have previously been measured in man, no crossovers between Hp, Tat, and Ctrb were observed in over 100 meioses in the mouse.

Esterase XXVII. Purification and characterization of esterase-9A of mouse kidney

Esterase-9A, which appears electrophoretically as a triplet of the bands III-50, III-40 and III-30, was isolated from the kidneys of male NMRI-mice by isoelectrofocusing and refocusing followed by repeated molecular sieve chromography. The overall purification was approx. 250 fold and each of the three bands was isolated separately. The band of the triplet nearest to the cathode, III-50, changed in vitro into the satellite bands III-40 and III-30 and, further, into the band III-22 not observed before in the homogenate. It is assumed that the band III-50 represents the original gene product. The molecular weight (45 000) of the band III-50 is identical with those of III-40 and III-30, as measured by analytical electrophoresis, whereas the molecular weight obtained by thin-layer chromatography was 51 000. There were no obvious signs that esterase-9 was composed of subunits. The Km constant for 4-nitrophenyl proprionate was identical for each of three bands. The esterase-9A is the first testosterone-dependent isozyme of the mouse carboxylesterase (carboxylicester hydrolase, EC 3.1.1.1) system which has been isolated.

Identity of esterase-22 and egasyn, the protein which complexes with microsomal β-glucuronidase

Recent experiments have demonstrated that egasyn not only sequesters β-glucuronidase in microsomes by forming high molecular weight complexes with β-glucuronidase, but also has carboxyl esterase activity. We have found several new phenotypes of egasyn-esterase after electrophoresis and isoelectric focusing of liver homogenates and purified egasyn of inbred and wild mouse strains. Several phenotypes corresponded in relative mobility and relative isoelectric point among inbred strains to that recently reported for esterase-22 by Eisenhardt and von Deimling [(1982). Comp. Biochem. Physiol. 73B:719]. This genetic evidence, plus a wide variety of comparative biochemical and physiological data, indicates that egasyn is identical to esterase-22. Both parental types of egasyn isozymes are expressed in heterozygous F1 progeny, suggesting that alterations in the egasyn structural gene are responsible for the altered isoelectric points. Also, egasyn is a monomer since no new esterase bands appear in F1 progeny. The variants in isoelectric point of egasyn map at or near the egasyn (Eg) gene within the esterases of cluster 1 near Es-9 on chromosome 8.