The aortic arch baroreceptor response to static and dynamic stretches in an isolated aorta-depressor nerve preparation of cats in vitro.

1. The aortic arch baroreceptors of cats were studied in an isolated aortic arch-depressor nerve preparation in vitro to analyse their transmission properties. 2. Sinusoidal length changes of varying amplitudes (0-34-1-2 mm) and frequencies (1-10 Hz) at a given pre-stretch were imposed on the isolated receptor zone to quantitate the velocity sensitivity of these receptors and to test the linearity of the system. 3. The receptor response was evaluated from the spike activity of single fibres of the depressor nerve in number of spikes per stimulus period, average discharge rate, instantaneous frequency, and phase angle between forcing function and instantaneous frequency. 4. The static response is characterized by a threshold, saturation range, and a relatively large linear part between these two non-linearities. 5. The aortic receptors exhibit rate sensitivity. Depending on the stimulus amplitude and frequency the phase angles between the forcing function and the instantaneous frequency ranged from -14 to -68 degrees. 6. The average discharge rate (spikes/sec) is sensitive to stimulus amplitude but not to stimulus frequency at near-threshold operation; in the linear part of the static response curve, the receptor response becomes independent of the stimulus mode. 7. The baroreceptors of the aorta share many properties with other mechanoreceptors of different species and organ systems, particularly with those of the rest of the circulation. They exhibit static and to some degree rate sensitivity and operate physiologically near their thresholds, i.e. the afferent arc of the baroreflexes is highly non-linear.


Fusion of host cell secondary lysosomes with the parasitophorous vacuoles of Leishmania mexicana-infected macrophages.

Secondary lysosomes of cultured mouse peritoneal macrophages were labeled with the electron-dense colloid saccharated iron oxide; the identity of the labeled structures was checked by the Gomori reaction for acid phosphatase. Amastigotes of Leishmania mexicana mexicana derived from mouse lesions were used to infect these macrophages in vitro. In electron micrographs of thin sections of infected macrophages the labeled secondary lysosomes were seen fused with the parasitophorous vacuoles without preventing subsequent multiplication of the parasites. A similar fusion probably occurs in vivo, and may provide a pathway through which not only nutrients but also drugs and host antibodies could reach the intracellular parasite.


Changes in lipase and phosphatase activities of rat spermatozoa in transit from the caput to the cauda epididymidis.

Rat spermatozoa from the cauda epididymidis, freed from their cytoplasmic droplets and acrosomes, were found to have a lower lipid content and to incorporate [14C]glucose into their glycerides and glycerophosphatides at a lower rate than spermatozoa from the caput epididymidis. Against the background of the activities of some glycolytic enzymes which remained constant the activity of alkaline phosphatase decreased in spermatozoa migrating through the epididymis, whereas the activity of monoglyceride lipase increased. The corresponding enzyme activities of non-flagellate germ cells of the testis were measured for comparison. The triglyceride lipase of non-flagellate germ cells and of spermatozoa from both caput and cauda epididymidis was activated by cyclic 3':5'-AMP.


Synthesis and evaluation of (Des-Asp1)angiotensin I as a precursor for (Des-Asp1)angiotensin II ("Angiotensin III").

The nonapeptide [des-Asp1]angiotensin I (IV), synthesized by Merrifield's solid-phase procedure, was tested as a possible substrate for the converting enzymes from porcine lung and plasma. IV, [des-Asp1]angiotensin II (III), [des-(Asp1,Arg2)]angiotensin II (V), [des-(Asp1,Arg2,Val3)]angiotensin II (VI), [Sar1,Ile8]angiotensin II (VII), and [des-Asp1,Ile8]angiotensin II (VIII) possessed 0.5, 20, 2, 0 less than 0.1, and less than 0.01% of the inotropic activity (rabbit atria), 1, 15, 5, 0, 3, and 0% secretory activity of the cat adrenal medulla, and 0.0, 150, 0.5, 3, and 10% of the adrenal steroidogenic activity of angiotensin II, respectively. When tested for their antagonistic activity in the above tissues, only VII and VIII were found to inhibit responses to angiotensin II. The pA2 values for VII and VIII were 8.31 and 10.0 in the adrenal cortex and 9.31 and 9.16 in the adrenal medulla, respectively. All these peptides were also tested as product inhibitors for the plasma and lung converting enzymes. With the plasma enzyme, the ID50 values were II, 1.6 X 10(-4) M; III, 5 X 10(-5) M; V, 1.2 X 10(-4) M; VI 5 X 10(-4) M; VII 5 X 10(-5) M; VIII, 5 X 10(-4) M. Thus, IV is a good substrate for converting enzymes from lung and plasma while all other compounds were inhibitors of these enzymes. The most potent inhibitors of converting enzyme were III followed by VII and VIII. With the exception of II and III, all the other analogs had very low intrinsic activities, per se. These results suggest (a) an alternate pathway for the formation of heptapeptide III, viz., by the action of converting enzyme on the nonapeptide IV, and (b) that III may also be acting as inhibitor of the converting enzyme by the feedback mechanism.


Synthesis of 5-amino-1-(5-deoxy-beta-D-ribofuranosyl)imidazole-4-carboxamide and related 5'-deoxyimidazole ribonucleosides.

5-Amino-1-(beta-D-ribofuranosyl)imidazole-4-carboxamide (1, AICA ribonucleoside) was converted in two steps to 5-amino-1-(5-deoxy-5-iodo-2,3-O-isopropylidene-beta-D-ribofuranosyl)imidazole-4-carboxamide (3) which was hydrogenated in the presence of Pd/C to yield 5-amino-1-(5-deoxy-2,3-O-isopropylidene-beta-D-ribofuranosyl)imidazole-4-carboxamide (4). The dehydration of 4 yielded 5-amino-1-(5-deoxy-2,3-O-isopropylidene-beta-D-ribofuranosyl)imidazole-4-carbonitrile (7). The compounds 3, 4, and 7 were deblocked with formic acid to furnish 5-amino-1-(5-deoxy-5-iodo-beta-D-ribofuranosyl)imidazole-4-carboxamide (6). 5-amino-1-(5-deoxy-beta-D-ribofuranosyl)imidazole-4-carboxamide (5), and 5-amino-1-(5-deoxy-beta-D-ribofuranosyl)imidazole-4-carbonitrile (8), respectively. Compound 8 was acetylated and then deaminated to give 1-(2,3-di-O-acetyl-5-deoxy-beta-D-ribofuranosyl)imidazole-4-carbonitrile (11). The compounds 8 and 11 were converted into 5-amino-1-(5-deoxy-beta-D-ribofuranosyl)imidazole-4-thiocarboxamide (9) and 1-(5-deoxy-beta-D-ribofuranosyl)imidazole-4-thiocarboxamide (12), respectively. The synthesis of 1-(5-deoxy-beta-D-ribofuranosyl)imidazole-4-carboxamide (13) was achieved for the first time by the treatment of 11 with hydrogen peroxide in the presence of ammonium hydroxide. The compounds were tested for antibacterial, antifungal, and antiviral activity, with 5 and 6 significantly inhibitory to Staphylococcus aureus.


Regulation of cellular calcium metabolism and calcium transport by calcitonin.

Calcitonin was studied in isolated kidney cells and in isolated mitochondria. A concentration of 10 ng/ml of synthetic calcitonin increases the cellular accumulation of 45Ca and the total cell calcium. The mitochondrial pool is increased several-fold. Kinetic analysis of the data shows that although the total cellular exchangeable calcium pool is enlarged, calcium influx and efflux are significantly depressed by calcitonin. The absence of phosphate or the presence of inhibitors of mitochondrial calcium transport completely abolish the effects of the hormone. In isolated mitochondria, the hormone stimulates the active calcium uptake and depresses the extramitochondrial calcium activity. Calcitonin counteracts the effects of cyclic AMP which stimulates the release of calcium from mitochondria and increases the extramitochondrial calcium activity. These data indicate that cellular calcium homeostasis is controlled by the mitochondrial calcium turnover. They suggest that calcitomin regulates the cell calcium metabolism and inhibits the transcellular calcium transport by stimulating the rate of calcium uptake by mitochondria which depresses cytoplasmic calcium activity.


Mechanism of inhibition of the proximal tubular isotonic fluid absorption by polylysine and other cationic polyamino acids.

The present study was initiated with the hope of clarifying the role of negative charges in the luminal brush border membrane in the overall process of trans-epithelial isotonic sodium and water absorption. Using micropuncture techniques, cationic polyamino acids such as polylysine (mol wt 100,000, 17,000 and 1,500-5,000, 1 mg/ml), tetralysine, polyornithine (mol wt 100,000, 1mg/ml), polyethyleneimine (2 mg/ml), polymyxin B (2 mg/ml), protamine sulfate (25 mg/ml) and histone (0.5 mg/ml) were perfused through the segments of rat kidney proximal tubule for 30 sec to 2 min. The rate of isotonic fluid absorption was measured before and after each perfusion with the Gertz's split drop method using Ringer's solution as a shrinking drop. Polylysine 100,000 and 17,000 and polyornithine were the most potent, inhibiting isotonic reabsorption by 93%. The sequence of inhibitory effect was: polylysine 100,000 congruent to polyornithine 100,000 congruent to polylysine 17,000 greater than polyethyleneimine greater than polylysine 1,500-5,000 congruent to polymyxin B greater than protamine sulfate congruent to histone. In contrast, tetralysine (2 mg/ml) showed no inhibitory effect. Electrical potential difference (p.d.) of the proximal tubular cells was destroyed within 10 sec of luminal perfusion with polylysine 100,000 (1 mg/ml). Simultaneously with the drop in p.d., electrical resistance of the luminal brush border membrane was nearly totally eliminated, whereas transepithelial input resistance remained unaltered. Furthermore, trypan blue dye was taken up by polylysine 100,000-perfused tubular cells but not by normal cells. Expanding drop analysis (mannitol solution as a split drop) was performed as a screening test to examine if the permeability for water and sodium in the lateral paracellular pathway is altered by polylysine 100,000. No significant difference was observed in the velocity of split drop expansion between untreated and polylysine-perfused tubules. A lower concentration of polylysine 100,000 (0.1 mg/ml) showed a much less inhibitory effect on fluid absorption and on cell p.d. These observations indicate that the strong inhibition on proximal tubular fluid absorption exerted by polylysine and perhaps also by other cationic polyamino acids is due not to modification of membrane negative charges but to the lysis of tubular cells by these polycations.


Irreversible stimulation of adenylate cyclase activity of fat cell membranes of phosphoramidate and phosphonate analogs of GTP.

The ability of 5'-guanylylimidodiphosphate (Gpp(NH)p) to stimulate irreversibly the adenylate cyclease activity of fat cell membranes has been studied by preincubating the membranes with this or related analogs followed by assaying after thoroughly washing the membranes. Activation can occur in a simple Tris-HCl buffer, in the absence of added divalent cations and in the presence of EDTA. Dithiothreitol enhances the apparent degree of activation, perhaps by stabilization. The importance of utilizing optimal conditions for stabilizing enzyme activity, and of measuring the simultaneous changes in the control enzyme, is illustrated. The organomercurial, p-aminophenylmercuric acetate, inhibits profoundly the activity of the native as well as the Gpp(NH)p-stimulated adenylate cyclase, but in both cases subsequent exposure to dithiothreitol restores fully the original enzyme activity. However, the mercurial-inactivated enzyme does not react with Gpp(NP)p, as evidenced by the subsequent restoration of only the control enzyme activity upon exposure to dithiothreitol. Thus, reaction with Gpp(NH)p requires intact sulfhydryl groups, but the activated state is not irreversibly destroyed by the inactivation caused by sulfhydryl blockade. GTP and, less effectively, GDP and ATP inhibit activation by Gpp(NH)p, but interpretations are complicated by the facts that this inhibition is overcome with time and that GTP and ATP can protect potently from spontaneous inactivation. These two nucleotides can be used in the Gpp(NH)p preincubation to stabilize the enzyme. The Gpp(NH)p-activated enzyme cannot be reversed spontaneously during prolonged incubation at 30 degrees C in the absence or presence of GTP, ATP, MgCl2, glycine, dithiothreitol, NaF or EDTA. The strong nucleophile, neutral hydroxylamine, decreases the Gpp(NH)p-activated enzyme activity and no subsequent activation is detected upon re-exposure to the nucleotide.


Mechanism of activation of adenylate cyclase by Vibrio cholerae enterotoxin. Relations to the mode of activation by hormones.

The influence of Vibrio cholerae enterotoxin (choleragen) on the response of adenylate cyclase to hormones and GTP, and on the binding of 125I-labeled glucagon to membranes, has been examined primarily in rat adipocytes, but also in guinea pig ileal mucosa and rat liver. Incubation of fat cells with choleragen converts adenylate cyclase to a GTP-responsive state; (-)-isoproterenol has a similar effect when added directly to membranes. Choleragen also increases by two- to fivefold the apparent affinity of (-)-isoproterenol, ACTH, glucagon, and vasoactive intestinal polypeptide for the activation of adenylate cyclase. This effect on vasoactive intestinal polypeptide action is also seen with the enzyme of guinea pig ileal mucosa; the toxin-induced sensitivity to VIP may be relevant in the pathogenesis of cholera diarrhea. The apparent affinity of binding of 125I-labeled glucagon is increased about 1.5- to twofold in choleragen-treated liver and fat cell membranes. The effects of choleragen on the response of adenylate cyclase to hormones are independent of protein synthesis, and they are not simply a consequence to protracted stimulation of the enzyme in vivo or during preparation of the membranes. Activation of cyclase in rat erythrocytes by choleragen is not impaired by agents which disrupt microtubules or microfilaments, and it is still observed in cultured fibroblasts after completely suppressing protein synthesis with diphtheria toxin. Choleragen does not interact directly with hormone receptor sites. Simple occupation of the choleragen binding sites with the analog, choleragenoid, does not lead to any of the biological effects of the toxin.


Distribution of parathyroid hormone-stimulated adenylate cyclase in plasma membranes of cells of the kidney cortex.

Free flow electrophoresis was employed to separate renal cortical plasma membranes into luminal (brush border microvilli) and contraluminal (basal-lateral membrane) fractions. During the separation adenylate cyclase activity was found to parallel the activity of Na+-K+-activated ATPase, an enzyme which is present in contraluminal but not in luminal membranes. In the basal-lateral membrane fraction the specific activities of adenylate cyclase and Na+-K+-activated ATPase were 4.4 and 4.6 times greater, respectively, than in the brush border fraction. The adenylate cyclase of the basal-lateral membrane fraction was specifically stimulated by parathyroid hormone which maximally increased enzyme activity eightfold. The biologically active (1-34) peptide fragment of paratyhroid hormone produced a 350% increase in adenylate cyclase activity. In contrast, calcitonin, epinephrine and vasopressin maximally stimulated the enzyme by only 55, 35 and 30%, respectively. These results indicate that adenylate cyclase, specifically stimulated by parathyroid hormone, is distributed preferentially in the contraluminal region of the plasma membrane of renal cortical epithelial cells.


Distribution of membrane-bound cyclic AMP-dependent protein kinase in plasma membranes of cells of the kidney cortex.

Renal cortical plasms membranes were separated by free flow electrophoresis into luminal (brush border microvilli) and contraluminal (basal-lateral membrane) fractions. These membranes were found to contain an intrinsic, self-phosphorylating system which consists of a cyclic AMP-dependent protein kinase, a phosphorprotein phosphatase and the substrate(s) of these enzymes. The kinase, but not the phosphatase, was stimulated by cyclic AMP; maximal (1.7-fold) stimulation was effected at a cyclic AMP concentration of 0.1 muM. The degree of phosphorylation of the brush borders was six times greater than that of the basal-lateral membranes in the absence of cyclic AMP and 2.3-fold greater in the presence of cyclic AMP. This preferential phosphorylation of the luminal membrane by membrane-associated protein kinase(s) may play a role in the parathyroid hormone-mediated alterations of solute reabsorption in the proximal tubule.


Isolation of RNA transcripts from the entire Sendai viral genome.

Three classes of viral transcripts (18S, 24S, and 33S) were isolated from viral ribonucleoproteins in Sendai virus-infected cells. Hybridization studies with virion minus strand genome RNA demonstrated that the 18S RNA contained transcripts from 60% of the viral genome while the 33S RNA contained transcripts from the entire viral genome. Brief heat of ME2SO treatment of the 33S RNA demonstrated that this RNA was composed of two classes: RNA which continued to sediment at 33S (33S RNA) and 18S RNA aggregates (18S RNA). The 33S RNA was determined to be a transcript from the 40% of the viral genome not protected by the 18S RNA. The aggregated 18S RNA does not appear to be an artifact of isolation.


Homology between avian oncornavirus RNAs and DNA from several avian species.

3H-labeled 35S RNA from avian myeloblastosis virus (AMV), Rous associated virus (RAV)-0, RAV-60, RAV-61, RAV-2, or B-77(w) was hybridized with an excess of cellular DNA from different avian species, i.e., normal or leukemic chickens, normal pheasants, turkeys, Japanese quails, or ducks. Approximately two to three copies of endogenous viral DNA were estimated to be present per diploid of normal chicken cell genome. In leukemic chicken myeloblasts induced by AMV, the number of viral sequences appeared to have doubled. The hybrids formed between viral RNA and DNA from leukemic chicken cells melted with a Tm 1 to 6 C higher than that of hybrids formed between viral RNA and normal chicken cell DNA. All of the viral RNAs tested, except RAV-61, hybridized the most with DNA from AMV-infected chicken cells, followed by DNA from normal chicken cells, and then pheasant DNA. RAV-61 RNA hybridized maximally (39%) with pheasant DNA, followed by DNA from leukemic (34%), and then normal (29%) chicken cells. All viral RNAs tested hybridized little with Japanese quail DNA (2 to 5%), turkey DNA (2 to 4%), or duck DNA (1%). DNA from normal chicken cells contained only 60 to 70% of the RAV-60 genetic information, and normal pheasant cells lacked some RAV-61 DNA sequences. RAV-60 and RAV-61 genomes were more homologous to the RAV-0 genome than to the genome of RAV-2, AMV, or B-77(s). RAV-60 and RAV-61 appear to be recombinants between endogenous and exogenous viruses.


Replication of picornaviruses. I. Evidence from in vitro RNA synthesis that poly(A) of the poliovirus genome is genetically coded.

A crude replication complex has been isolated from poliovirus-infected HeLa cells and used for synthesis of poliovirus replicative intermediate (RI) RNA, replicative form (RF) RNA, and single-stranded (SS) RNA in vitro. All three classes of virus-specific RNA synthesized in vitro are shown to contain poly(A). Poly(A) of RF and of SS RNA [RF-poly(A) and SS-poly(A)] has a chain length (50 to 70 nucleotides) that is shorter than that of poly(A) of in vivo-synthesized RNAs. Poly(A) of RI [RI-poly(A),] however, is at least 200 nucleotides long and, therefore, larger than poly(A) of RI isolated from HeLa cells 4 h after infection. The crude membrane-bound replication complex contains a terminal adenylate transferase activity that is stimulated by Mn2+ and the addition of an (Ap)2AOH primer. This transferase activity is found also in extracts of mock-infected cells. Partial purificaiton of the replication complex in a stepwise sucrose gradient, in which the viral replicase is associated with the smooth cytoplasmic membrane fraction, does not remove the terminal transferase. However, when the partially purified replication complex is treated with deoxycholate and sedimented through a sucrose gradient, a soluble replication complex can be isolated that is free from terminal adenylate transferase. This soluble replication complex was found to synthesize viral RNA-linked poly(A) longer in chain length than that synthesized by the crude replication complex. Taking into account the 5'-terminal poly(U) in poliovirus minus strands, our data suggest that polyadenylation of poliovirus RNA occurs by transcription and not by end addition. When compared to other viral systems, poliovirus and, probably, all picornaviruses appear to be unique in that the poly(A) of their genome is genetically coded.


Human cytomegalovirus. IV. Specific inhibition of virus-induced DNA polymerase activity and viral DNA replication by phosphonoacetic acid.

Phosphonoacetic acid specifically inhibited human cytomegalovirus DNA synthesis in virus-infected human fibroblasts as detected by virus-specific nucleic acid hybridization. Inhibition was reversible; viral DNA synthesis resumed upon the removal of the drug. The compound partially inhibited DNA synthesis of host cells in the log phase of growth but had little effect on confluent cells. Studies of partially purified enzymes indicated that phosphonoacetic acid specifically inhibited virus-induced DNA polymerase and had only a slight effect on normal host cell enzymes. The drug was shown to interact directly with virus-induced enzyme but not with the template-primers.


Synthesis of herpes simplex virus, vaccinia virus, and adenovirus DNA in isolated HeLa cell nuclei. I. Effect of viral-specific antisera and phosphonoacetic acid.

Purified nuclei, isolated from appropriately infected HeLa cells, are shown to synthesize large amounts of either herpes simplex virus (HSV) or vaccinia virus DNA in vitro. The rate of synthesis of DNA by nuclei from infected cells is up to 30 times higher than the synthesis of host DNA in vitro by nuclei isolated from uninfected HeLa cells. Thus HSV nuclei obtained from HSV-infected cells make DNA in vitro at a rate comparable to that seen in the intact, infected cell. Molecular hybridization studies showed that 80% of the DNA sequences synthesized in vitro by nuclei from herpesvirus-infected cells are herpesvirus specific. Vaccinia virus nuclei from vaccinia virus-infected cells, also produce comparable percentages of vaccinia virus-specific DNA sequences. Adenovirus nuclei from adenovirus 2-infected HeLa cells, which also synthesize viral DNA in vitro, have been included in this study. Synthesis of DNA by HSV or vaccinia virus nuclei is markedly inhibited by the corresponding viral-specific antisera. These antisera inhibit in a similar fashion the purified herpesvirus-induced or vaccinia virus-induced DNA polymerase isolated from infected cells. Phosphonoacetic acid, reported to be a specific inhibitor of herpesvirus formation and the herpesvirus-induced DNA polymerase, is equally effective as an inhibitor of HSV DNA synthesis in isolated nuclei in vitro. However, we also find phosphonoacetic acid to be an effective inhibitor of vaccinia virus nuclear DNA synthesis and the purified vaccinia virus-induced DNA polymerase. In addition, this compound shows significant inhibition of DNA synthesis in isolated nuclei obtained from adenovirus-infected or uninfected cells and is a potent inhibitor of HeLa cell DNA polymerase alpha.


Effect of the Fv-1 locus on the titration of murine leukemia viruses.

Titration of N- and B-tropic murine leukemia viruses on sensitive and resistant cell lines has been studied by direct XC plaque assay and infective center assay. The titration of cloned B-tropic virus by infective center assay on BALB/3T3 (Fv-1b/b) and NIH/3T3 (Fv-1n/n) cells gave one-hit patterns, with 100-fold less infected NIH/3T3 cells than BALB/3T3 cells. The titration of B-tropic virus on DBA/2 cells (Fv-1n/n) was also a one-hit. The titration of a one-hit curve, and there were about 100-fold less infected BALB/3T3 cells than NIH/3T3 cells. Comparable results were obtained by titrating the cloned N-tropic virus on congenic SIM (Fv-1n/n) and SIM.R (Fv-1b/b) cells or the Gross N-tropic virus on BALB/3T3 cells. Therefore, our data indicate that the multiple-hit phenomenon described previously may not be an essential part of the Fv-1 gene restriction.


tRNA's associated with the 70S RNA of avian myeloblastosis virus.

The distribtuion of various amino acid tRNA's in the 4S RNA components of avian myeloblastosis virus (AMV) and in 4S RNA prepared from chicken cmbryo cells, chicken myeloblasts, and chicken livers was determined. This was done by aminoacylating the 4S RNA samples with a mixture of 17 radioactive amino acids and subsequently identifying the tRNA-accepted amino acids on an amino acid analyzer after deacylation. In embryo cells, myeloblasts, and liver, tRNA's accepting all 1m amino acids were demonstrated. "Free" AMV 4S RNA was characterized by very low quantities of glutamate, valine, and tyrosine tRNA's. RNAs accepting all 17 amino acids, with the exception of tyrosine, were shown to be present in the "70S-associated" 4S RNA which dissociates at 60 C. The bulk of the 70S-associated 4S RNA was dissociated at 60 C at low ionic strength with a concomitant conversion of 70S RNA to 35S RNA. The residual associated 4S RNA was dissociated by further heating of the 35S RNA to 80 C; tryptophan tRNA accounted for greater than 90% of the total amino acid accepting activity in this fraction. The results support other studies in suggesting that tryptophan tRNA may serve as a primer for DNA synthesis in AMV, as has been shown in Rous sarcoma virus.


Block to multiplication of adenovirus serotype 2 in monkey cells.

The block to adenovirus 2 (Ad2) multiplication in monkey cells can be overcome by coinfection with simian virus 40 (SV40). To identify this block we have compared the synthesis of Ad2 proteins in monkey cells infected with Ad2 alone (unenhanced) or with Ad2 plus SV40 (enhanced). Synthesis of viral proteins in enhanced cells was virtually identical to that found for permissive infection of human cells by Ad2 alone. In contrast, the unenhanced cells were strikingly deficient in the production of the IV (fiber) and 11.5K proteins whereas the synthesis of 100K and IVa2 was normal. Synthesis of a number of other proteins such as II, V, and P-VII was partially reduced. A similar specific reduction in synthesis of these proteins was found when their messages were assayed by cell-free translation. This result suggests that the block to Ad2 protein synthesis is at the RNA level rather than with the translational machinery of monkey cells. Analysis of the complexity and the concentration of Ak2-specific RNAs, using hybridization of restriction endonuclease fragments of the Ad2 genome to increasing concentrations of RNA, shows that although all species of late Ad2 mRNA are present, the concentration of several species is reduced sevenfold or more in unenhanced monkey cells as compared with enhanced cells. These species come from regions of the genome known to encode the deficient proteins. A model for the failure of adenovirus to multiply in monkey cells, based on abnormal processing of specific adenovirus messages, is presented.


Hepatitis in skunks caused by the virus of infectious canine hepatitis.

Two cases of acute, fatal, hepatitis occurred in young, striped skunks (Mephitis mephitis) trapped in southern Ontario. Histologically, lesions in the liver were similar to infectious canine hepatitis. A virus was isolated which produced large intranuclear inclusions in dog kidney cell cultures. These inclusions were Feulgen-positive and fluoresced green with acridine orange stain. The skunk hepatitis isolate was identified as the virus of infectious canine hepatitis by virus neutralization tests.


Histopathological observations of spontaneous regression of Rous sarcomas in Japanese quails.

Turmors induced in Japanese quails by the Schmidt-Ruppin strain of Rous sarcoma virus were examined histopathologically. The following three phases were recognized in the quails whose tumors regressed finally (regressor). Phase I was between days 4 and 7 of virus inoculation, when growth of tumor cells was seen with predominant infiltration of heterophils. Phase II, from days 10 to 14, was characterized by necrosis of tumor cells and focal accumulation of lymphoid cells which frequently formed follicle-like nodules. In phase III from days 18 to 24, tumor cells and heterophils disappeared, whereas diffuse infilitration of lymphoid cells, plasma cells and histiocytes were demonstrated. In the quails whose tumors progressed (progressor), growth of tumor cells and infiltration of heterophils at phase I seemed to follow a pattern similar to that of regressors, but subsequent infiltration and focal accumulation of lymphoid cells were rare. These morphological findings suggested an immunological reaction against tumor cells by lymphoid cells.


Characterization of "Pasteurella" piscicida isolated from white perch and cultivated yellowtail.

The morphological, physiological, and biochemical characteristics of twelve "Pasteurella" piscicida strains isolated from white perch and yellowtail are described and the present uncertain taxonomic status of the organisms is discussed. The organisms isolated were gram-negative rods showing bipolar staining and pleomorphism. No spores or flagella were observed. They were non motile and viscid colonies were formed. Growth was observed in a temperature range of 20 to 30 C and the salinity range of growth was between 0.5% and 4.0%. They were aerobic and facultative anaerobic. Oxidase and catalase were produced. Nitrates were not reduced to nitrites. Lysine and ornithine decarboxylases were not produced but arginine dihydrolase was produced. The egg yolk reaction was positive. Tween 80 and tributyrin were decomposed. Phosphatase was produced. Beta hemolysis was revealed on a medium containing defibrinated blood from chickens or carp but not from mammals. Methyl red and Voges-Proskauer tests were positive, and acetoin was produced from 2,3-butanediol. Glucose was fermented without gas production. Acid was produced from fructose, mannose, galactose, sucrose, maltose, dextrin and glycerol. These organisms differed from all other members of the genus Pasteurella with respect to nitrate reduction, arginine dihydrolase, Voges-Proskauer and methyl red tests. The formation of viscid colonies and inability to grow in a medium without sodium chloride or at 37 C were additional characteristics of these organisms.


Control of plasma aldosterone in normal man during upright posture.

Plasma aldosterone, plasma renin activity (PRA), plasma cortisol as parameter of ACTH activity and the serum concentrations of sodium and potassium were determined at short time intervals in 10 healthy students after an overnight bedrest and during three hours of ambulation. While PRA rose significantly within 15 minutes of orthostasis in all students, plasma aldosterone showed a similar rapid increase in some of the subjects only. These persons demonstrated also a simultaneous increase of serum potassium or of plasma cortisol. Plasma aldosterone rose not before 30 to 60 minutes after change to the upright position in subjects who showed neigher plasma cortisol nor serum potassium increases. It is concluded that the immediate rise of plasma aldosterone during orthostasis seems to depend on a stimulation by ACTH of by potassium. The main stimulus of plasma aldosterone during orthostasis appears to be the renin angiotensin system. If the aldosterone response to posture is mediated only through this system a delay of 30 to 60 minutes is observed.


Ultrastructure and virus particles of L1210/Mes. A hamster-adapted murine leukemia.

The ultrastructural features and virus particle content of L1210/Mes., the hamster-adapted strain of murine leukemia L1210, has been studied in cheek pouch tumors and cell cultures. In comparison with the parental murine tumor in culture, L1210/Mes. has experienced certain morphologic alterations, such as a striking increase in cell size, enlarged and more bizarre nuclei, and an increase in the number of lysosome-related structures. Type A (intracytoplasmic and intracisternal) and type C virus particles could be demonstrated in cell cultures of murine leukemia L1210, and in cheek pouch grafts and cell cultures of L1210/Mes. No. "H" (or "R") hamster-associated virus particles, however, could be distinguished in L1210/Mes. tumors or in vitro cultivated cells. It appears that a murine lymphoma capable of serial propagation in a xenogeneic host permits the replication of virus particles structurally indistinguishable from those indigenous to it.


Electron paramagnetic resonance and nanosecond fluorescence depolarization studies on creatine-phosphokinase interaction with myosin and its fragments.

Recent reports in the literature have indicated a physical association of creatine-phosphokinase (CPK) with the tail portion of the myosin molecule. The present paper describes further studies on the interaction of CPK with myosin and myosin fragments, using the techniques of electron paramagnetic resonance (EPR) and nanosecond fluorescence depolarization. From EPR work, spin-labeled CPK appears to interact with myosin, tail-less myosin (heavy meromyosin [HMM]), and myosin heads (subfragment-1 [S1]), the extent of interaction being proportional to the S1 content of myosin or its fragments. Spin-labeled CPK did not evidence interaction with the headless myosin "rods," with myosin tails (light meromyosin [LMM]), with S2 necks (which connect S1 to the rest of the myosin molecule), or with actin. When a fluorescent dye is directed to the essential epsilon-amino group of CPK, nanosecond fluorescence depolarization studies indicate a substantial interaction with myosin, HMM, and S1, but very little with F-actin. When the "fast-reacting" thiol of the S1 moiety or the "essential thiol" of CPK was labeled with either a fluorescent dye or a spin label, no interaction between CPK and myosin (or S1) was detected.


Birefringence changes in vertebrate striated muscle.

The changes in birefringence in the rigor to relax transition of single Triton-extracted rabbit psoas muscle fibers have been investigated. The total birefringence of rigor muscle fibers was dependent on sarcomere length and ranged from (1.46 +/- 0.08) X 10(-3) to (1.60 +/- 0.06) X 10(-3) at sarcomere lengths from 2.70 mum to 3.40 mum. An increase in total birefringence was measured dependent on sarcomere length when 55 single fibers were relaxed from the rigor state with Mg-ATP. Pyrophosphate relaxation produced a smaller increase in retardation when compared to Mg-ATP. The expected change in intrinsic birefringence during the rigor to relax transition was calculated assuming a hinge function of the subfragment 2 moiety of myosin. The changes in birefringence during isometric contraction and relaxation have been discussed in relation to possible structural changes.


Transient kinetic and isotopic tracer studies of the myosin adenosine triphosphatase reaction.

From transient kinetic studies of the Mg2+-dependent adenosine triphosphatase of myosin subfragment 1, prepared from rabbit skeletal muscle, a seven-step mechanism has been proposed. Features of this mechanism include two-step processes for ATP and ADP binding in which the binary complex isomerizes in addition to a rapid nucleotide association step. In the case of ATP a large negative standard free energy change is associated with the isomerization. An overall rate-limiting isomerization of the myosin-product complex prior to product release has been identified. Studies on the mechanism of cleavage of ATP bound to the active site indicate the process is readily reversible and can account for the observation that more than one oxygen  of the product phosphate arises from water. This proposal has been substantiated by the finding that the oxygen atoms of the gamma-phosphoryl group of bound ATP also undergo extensive exchange with water.


Intermediate complex of ATP hydrolysis and synthesis by muscle proteins.

Myosin catalyzed exchange between 32Pi and ATP in reaction medium during its enzymatic hydrolysis of ATP only by a very small amount. Addition of actin increased to a great extent the rate of incorporation of 32Pi in the presence of Mg. Glycerinated smooth muscle fibers also exhibited the ability to exchange 32Pi and ATP upon the application of external force (repeated stretching and releasing). A schematic mechanism of the action of actin and external force on acceleration of 32Pi incorporation is proposed and the importance of the M-ADP complex for force generation is suggested.


The quantitative measurement of rotational motion of the subfragment-1 region of myosin by saturation transfer epr spectroscopy.

According to current models of muscle contraction (Huxley, H. E., Science 164: 1356-1366 [1969]), motion of flexible myosin crossbridges is essential to the contractile cycle. Using a spin-label analog of iodoacetamide bound to the subfragment No. 1 (S1) region of myosin, we have obtained rotational correlation times (tau 2) for this region of the molecule with the ultimate goal of making quantitative measurements of the motion of the crossbridges under conditions comparable to those in living, contracting muscle. We used the newly developed technique of saturation transfer electron paramagnetic resonance spectroscopy (Hyde, J.S., and Thomas, D.D., Ann. N.Y. Acad Sci. 22:680-692 [1973]), which is uniquely sensitive to rotational motion in the range of 10(-7)-10(-3) sec. Our results indicate that the spin label is rigidly bound to S1 (tau 2 for isolated S1 is 2 X 10(-7) sec) and that the motion of the label reflects the motion of the S1 region of myosin. the value of tau 2 for the S1 segment of myosin is less than twice that for isolated S1, while the molecular weights differ by a factor of 4, indicating flexibility of myosin in agreement with the conclusions of Mendelson et al. (Biochemistry 12:2250-2255 [1973]). Adding F-actin increses tau 2 in either myosin or isolated S1 by a factor of mearly 103, indicating rigid immobilization of S1 by actin. Formation of myosin filaments (at an ionic strength of 0.15 or less) increses tau 2 by a factor of 10-30, depending on the ionic strength, indicating a decrease of the rotational mobility of S1 in these agregates. The remaining motion is at least a factor of 10 faster than would be expected for the filament itself, suggesting motion of the S1 region independent of the filament backbone but slower than in a single molecule. F-actin has a strong immobilizing effect on labeled S1 in myosin filaments (in 0.137 M KC1), but the immobilization is less complete than that observed when F-actin is added to labeled myosin monomers (in 0.5 M KC1). A spin-label analog of maleimide, attached to the SH-2 thiol groups of S1, is immobilized to a much lesser extent by F-actin than is the label on SH-1 groups. The maleimide label also was attached directly to F-actin and was sufficiently immobilized to suggest rigid binding to actin.


Lymphography in childhood: six years experience with 242 cases.

From January, 1969 - December, 1974, 242 children less than 15 years of age underwent lymphography at the National Cancer Institute, Milan. Successful lymphatic cannulation was accomplished in 97% (440/463) of the sites where it was attempted. No major or permanent complications were encountered, although minor untoward effects might not have been recorded. In those children undergoing biopsy of opacified lymph nodes, lymphographic-histologic correlation was 98% (45/46). Nonspecific reactive hyperplasia lymphographic patterns were encountered in 36% of all studies, confirming its high incidence in the pediatric age group. This study has shown that lymphography in childhood can be as readily performed as in the adult and that its diagnostic accuracy is acceptable. As in adults, it is useful in treatment planning, evaluating results of therapy, and detecting recurrent tumor. The frequent occurrence of nonspecific reactive hyperplasia in the pediatric lymphogram should not be mistaken for evidence of tumor, particularly lymphoma.


Endolymphatic radiotherapy in malignant lymphomas: its potential "prophylactic" value in cases with negative lymphograms.

From 1961 to 1969 426 patients (208 with Hodgkin's disease and 218 with non-Hodgkin's lymphoma) underwent endolympatic radiotherapy with Lipiodol 131I at the National Cancer Institute of Milano. For this study, only those patients with stage I, II, or III disease (with or without systemic symptoms), who were not previously treated, and who had a complete follow-up were reviewed. It appears that while in the cases where there is lymphographic evidence of involved lymph nodes, endolymphatic radiotherapy is not of value, in the cases with apparently negative lymphography, endolymphatic radiotherapy can reduce the incidence of relapse in the inguino-retroperitoneal nodes to a statistically significant degree.


Pain. A central inhibitory balance theory.

Knowledge of the segmental neuronal circuitry for noxious and non-noxious input is a necessary prelude to understanding pain mechanisms. Significant gaps in our understanding of the synaptology of these pathways in the dorsal horn have hindered progress in this area. This report describes the excitatory and inhibitory circuits that supply the marginal neurons of the spinal cord, whose role in nociceptive mechanisms is well  established. On the basis of these data, it is proposed that large primary afferents (non-noxious) provide heavy inhibitory input to marginal neurons via gelatinosa cells. Conversely, small primary afferents (nociceptive) provide excitatory input to marginal neurons but relatively little inhibitory feedback to these cells via gelatinosa neurons. The modulation of pain-producing input depends thus on the balance between large-fiber and small-fiber activity via a postsynaptic inhibitory mechanism acting on the nociceptive relay neurons. This theory accounts satisfactorily for the modulation of pain by counterirritation and by various methods of stimulating large-fiber input.


[The treatment of inoperable juvenile angiofibromas (author's transl)].

Demonstrating a case of juvenile angiofibroma penetrating the base of the scull and growing mainly endocranially we refer to Honvan (Fostestrolum) treatment of this kind of tumor. The effect of this treatment can be shown histologically as a distinct sclerosis of the tumor tissue. Even in long term application of this drug only very few side effects are found.


A possible effect of testosterone on the adenosine 3'-5' cyclic monophosphate levels in rat cerebral cortex. A brief note.

Based on a correlation of serum testosterone levels and cortical cyclic-AMP concentrations in a normal aging population of male CD Fischer rats and a demonstration of decreased cortical cyclic-AMP subsequent to injection of testosterone, it was concluded that a cause and effect relation between hormone concentrations and brain cyclic nucleotide levels is tenable.


Inhibition of hepatic lipogenesis by adenine nucleotides.

Incubation of liver slices and isolated liver cells with adenosine cyclic-3',5'-monophosphate at concentrations which inhibit lipogenesis was found to expand the pool size of the noncyclic adenine nucleotides in the intact cells of the preparations. This observation led to studies which demonstrated that adenosine and adenosine-5'-monophosphate also inhibited lipogenesis and expanded the adenine nucleotide pool size. It is proposed but not proven that the increase in intracellular nucleotides produced by adenosine-5'-monophosphate, adenosine cyclic-3',5'-monophosphate, and adenosine may have an adverse effect upon the synthesis of fatty acids. Because of the expansion of the adenine nucleotide pool size, high concentrations of adenosine cyclic-3',5'-monophosphate should not be used to investigate the mechanism responsible for hormonal regulation of lipogenesis. As an added complication, exogenous adenosine-5'-monophosphate was found to produce a small but significant increase in the intracellular concentration of adenosine cyclic-3',5'-monophosphate of isolated liver cells. This effect also may be a factor in the inhibition of lipogenesis by adenosine-5'-monophosphate. Low concentrations of N6, O2'-dibutyryl adenosine cyclic-3',5'-monophosphate were found to inhibit lipogenesis without increasing the intracellular adenine nucleotide content of either liver slices or isolated liver cells. It is concluded that studies on the mechanism of glucagon regulation of lipogenesis should be carried out with glucagon or low concentrations of N6, O2'-dibutyryl adenosine cyclic-3',5'-monophosphate.


Immunoassay of human plasma apolipoprotein B.

A specific and precise double antibody immunoassay for human plasma apolipoprotein B (apoB) was developed and applied in normolipidemic and hyperlipidemic subjects. The intra-assay coefficient of variation was ca. 9%. The distributions of total apoB and low density lipoprotein (LDL) apoB in a randomly selected, healthy, fasting population (n = 349) was slightly skewed with a mean total apoB of 81 mg/100 ml and LDL apoB of 72 mg/100 ml. The 90th percentile cutoffs for total apoB and LDL apoB were 106 and 97 mg/100 ml, respectively. Regarding total apoB, women showed a statistically significant increase (r = 0.463, p less than 0.001) with age (30-65) and an average annual increment of plasma apoB of 1.1 mg/100 ml. In contrast, men showed only a slight increase of apoB from the 4th to 5th decade, with an average annual increment of 0.7 mg/100 ml (r = 0.201, 0.02 less than p less than 0.05). Similarly, regarding LDL apoB, women showed an increase of 1.0 mg/100 ml/year from the 4th to 7th decade (r = 0.501, p less than 0.001), whereas men's LDL apoB did not increase significantly with age (r = 0.114, 0.2 less than p less than 0.3, for ages 30-49). Six of ten normal young subjects showed essentially no physiological variation in fasting apoB levels over a 10-wk period, whereas four had a variation of ca. 5% or less. LDL apoB represented ca. 90% of the total apoB in normolipidemic and type II plasma samples (86% in type IV samples) but only 68% in type III plasmas (n = 7). The ratios of LDL cholesterol-LDL apoB were similar for the random and hyperlipoproteinemic groups, ranging from a high of 1.8 for type IIa to a low of 1.5 for type IV. The ratio of cholesterol to apoB was significantly elevated (p less than 0.002) in the d less than 1.006 fraction of the type III plasma samples compared to the random and type II groups.


Experimental endocrinopathies.

Methods are described for maintaining hypophysectomized rats (model for panhypopituitarism). Prolactin-secreting pituitary tumors can be induced in rats or mice by administration of estrogens; thyroid stimulating hormone-secreting tumors will occur in some mice after thyroid ablation by radioactive iodine. Estrogens in hamsters usually produce intermediate lobe tumors of the pituitary associated with hypothalamic degeneration. Sex hormone-secreting adrenal tumors can follow surgical gonadectomy in mice. Spontaneous corticoid-secreting adrenal tumors may occur spontaneously in Osborne-Mendel rats. Secretory gonadal tumors have been induced by transplantation of a gonad into the spleen of a gonadectomized host. Both secretory and non-secretory ovarian tumors can be produced by irradiation or chemical carcinogens in mice. In some mice, secretory testicular tumors can be produced by estrogen administration. Thyroid tumors can be induced in rodents by various kinds of goitrogens and irradiation. Parathyroid hyperplasia may occur with spontaneous renal disease in rats. A syndrome simulating diabetes mellitus can occur in rare strains of mice or can be induced by chemical destruction of the islets of Langerhans with alloxan.


[Bronchial asthma. Pathogenetic assumptions - therapeutic guidelines - the significance of synthetic corticotrophin (ACTH) in its treatment (author's transl)].

The treatment procedure with 121 in-patients suffering from respiratory obstruction is described following a short review of the etiology and pathogenesis of bronchial asthma, and the consequences for treatment resulting therefrom. While patients with chronic bronchitis caused by an emphysema of the lungs were treated primarily with antibiotics, synthetic corticotrophin (Synacthen, Synacthen Depot) and/or cortisone were prescribed in the treatment of 3/4 of the cases involving asthmatics and patients with chronic asthmatoid bronchitis caused by an emphysema. In severe cases ACTH was combined with cortisone. As a result of corticotrophin's direct effect on the bronchial muscles and the mast cells, along with its stimulation of the adrenal cortex, treatment with ACTH has the advantage over the classic cortisone treatment of retaining the functional ability of the adrenal cortex and reducing the amount of cortisone needed. No ACTH-caused side effects were reported. Included are tables containing guidelines for therapy suitable in private practice.


[Respiratory tract infections of viral origin in infants. Examination of 103 serum pairs in autumn and following spring].

In autumn 1973 und about four months later in spring 1974 blood samples were taken from the same 103 children aged from a few months to five years and coming from different parts of Bavaria. We examined the samples for complement-fixing antibodies against the influenza virus types A, B and C, the parainfluenza virus types 1, 2 and 3, the adenoviruses and the respiratory-syncytial virus, and for neutralising antibodies against the ECHO virus type 25. At the commencement of the investigation the most frequently detected antibodies were those against adenoviruses (66.0%), ECHO virus type 25 (22.3%), influenza virus type A (15.5%) AND THE RESPIRATORY-SYNCYTIAL VIRUS (10.7%). Over autumn and winter in the four month interval between the drawing of blood samples the most frequently observed infections were those caused by the respiratory-syncytial virus (35.9%), the adenoviruses (13.6%) and the influenza viruses A and B (6.8%). By comparing the results of two blood tests from the same individual infections could be recognised more often than would have been the case if comparing the results of two different groups of children examined in autumn and spring.


A mutant of Aspergillus nidulans defective in NAD-linked glutamate dehydrogenase.

A mutation leading to partial loss of NAD-linked ("catabolic') glutamate dehydrogenase does not affect the regulation of ammonium-repressible activities in Aspergillus nidulans. This mutation has been used to show that NAD-linked glutamate dehydrogenase does not normally participate in ammonium assimilation. A mutation leading to loss of NADP-linked ("anabolic') glutamate dehydrogenase has been used to show that NADP-linked glutamate dehydrogenase is not normally involved in glutamate catabolism. Strains defective in either enzyme are useful for determining which amino acids are metabolised via transamination to yield glutamate rather than via deamination to yield ammonium.


The effect of triiodothyronine on myocardial protein kinases.

Rats treated with T3 (triiodothyronine) showed an increased heart weight after 3 days reaching 100% after 3 weeks of treatment compared to untreated controls. Cytosol protein kinases were not significantly different in the T3 treated rats compared to controls. The protein kinase activity of the NHP (nonhistone proteins) increased after 2 hours and doubled after 3 days for each substrate tested. After 1 week of T3 treatment the protein kinase activity returned to the control value and remained at the control level for the remainder of the 3 week experimental period. A study of the distribution of protein kinase activity in the NHP by disc gel electrophoresis showed that there was a difference in the distribution of some peaks in the T3 treated animals compared to the controls. T3 in concentrations from 10(-11) to 10(-3) M had no in vitro effect on the phosvitin kinase activity of NHP and of cytosol.


The involvement of superoxide anions in the autoxidation of various cofactors of cysteamine-oxygenase.

The reduction of tetranitroblue tetrazolium with cysteamine, mediated by a number of dyes, elemental sulphur, elemental selenium and selenide, under aerobic conditions, was inhibited to various extent upon addition of superoxide dismutase. A strict parallelism between the ability to produce O2- ions and the property of those compounds to act as cofactors for cysteamine-oxygenase, to yield hypotaurine, has been observed. Based on the fact that the autoxidation of cysteamine also gives rise to O2- formation, though to a minor extent, we propose a mechanism for cysteamine-oxygenase action. This mechanism was derived from the data obtained in the model system studied.


Glucocorticoid receptors and lymphocytolysis in normal and neoplastic lymphocytes.

When cells of the thymus or mouse leukemias P288 and L1210 are exposed in vitro to the potent synthetic glucocorticoid, 3H-Triamcinolone acetonide, the steroid enters the cells passively and binds to macromolecules in the cytoplasm. At 37 degrees C this hormone-receptor complex enters the nucleus and associates with the chromatin. The association with chromatin occurs not only in the corticosteroid-sensitive rat thymocytes and mouse tumors P288 and P1798S but also in the corticosteroid-resistant mouse tumors L1210 and P1798R. An apparent correlation, although not absolute, exists between the content of glucocorticoid-binding macromolecule and the sensitivity of the lymphocytes studied to the lytic effect of glucocorticoids; the sensitive cells having more receptor than the resistant cells. The process of lysis is attributed to the release from the much larger stores of triglyceride in thymus and sensitive lymphoma cells, of a large pool of FFA which causes focal damage to the nuclear membrane resulting in karyorrhexis and, subsequently, to cytolysis. Resistance is attributed to the capacity for preventing the accumulation of greater than about 0.5 fmole FFA/cell. Resistant cells induced to accumulate greater amounts, even for a few minutes, ultimately undergo lysis. Most effective in accomplishing this are branched chain fatty acids of C-8 and higher, which block FFA metabolism, causing accumulation which results in cytolysis.


A directed alteration of ribonuclease specificity. Hydrolysis of polyribonucleotides containing modified cytosine bases.

The action of ribonucleases on poly and oligoribonucleotides containing cytosine bases modified by methoxyamine and bisulphite was examined. Resistance of phosphodiester bonds in (Cp)nXp (where n greater than or equal to 1 and X stands for A, G or U) to T2 RNase hydrolysis was observed if substrates were modified chemically. The phenomenon formed the basis for isolation of (Cp)nXp blocks as an additional tool in sequence investigations. After modification of cytosine pancreatic RNase was unable to hydrolyse (Cp)nUp blocks. Therefore the specificity of pyrimidyl RNase may be narrowed to 'uridyl RNase'.


Influence of a specific histone kinase on the physico-chemical properties of chromatin in situ.

The influence of a specific histone kinase, phosphorylating lysine-rich histone F1, F2a2, F2b, on the physico-chemical properties of the chromatin in the whole undestroyed fixed cell, has been investigated. It was found that the exogenous histone kinase penetrates into the nuclei of the undestroyed fixed cells and into the isolated unfixed nuclei and changes the physico-chemical properties of the chromatin there, bringing about an increase in binding of a basic dye acridine orange and a decrease in its stability to heat.


[Results of non-specific iliac crest biopsy in patients with bronchial carcinoma (author's transl)].

Systematic bone marrow examinations in patients with bronchial carcinoma (n equals 75) gave positive findings (identification of tumor cells) in 12% related to all types of tumors and in 32% of microcellular carcinoma. The information value of this method is relatively small with regard to the assessment of operability, because positive findings in stages I and II are extremely rare. Non-specific iliac crest biopsy provides therapeutically and prognostically important pointers in patients who are to be irradiated (tumor stage III and all microcellular tumors of stages I-III), because a positive bone marrow finding must be valued as stage IV.


[Synthetic ACTH in theumatic diseases: (author's transl)].

44 patients with acute irritations of degenerative articular diseases and 66 patients with soft tissue rheumatism were treated with Synacthen Depot (tetracosactide hexacetate 1 mg/ml). During the first three days of treatment the patients received an average of 2 ampoulbs Synacthen Depot, then treatment was continued at 3-4 day intervals. In the degenerative joint disease group 86.4% and 89.4% in the group with soft tissue rheumatism became free of complaints or improved with the ACTH treatment. Most of the patients with degenerative joint diseases needed a 4 week treatment; 1/3 of the patients with soft tissue rheumatism were already free of complai


Chemical induction of presumed dominant-lethal mutations in postcopulation germ cells of mice. I. Relative sensitivity between pre- and postcopulation germ cells to isopropyl methanesulfonate.

Females from (C3H X 101)F1 and a mixed stock were injected intraperitoneally with either 25 or 50 mg/kg isopropyl methanesulfonate (IMS) or Hanks' solution 4-5 h after the midpoint of the dark period during which mating occurred. It was determined that at the time of treatment the great majority of oocytes were undergoing second meiotic division. For comparison, the same doses of IMS were given to females treated within 3-5 days prior to mating (predominantly dictyate oocytes) or to males treated within 4-5 days prior to mating (sperm in vas and epididymis). The frequencies of presumed dominant lethals induced by 50 mg/kg IMS in sperm treated in vas and epididymis, dictyate oocytes, and germ cells in mated females are 22%, 19%, and 79%, respectively, for (C3H X 101)F1 and 26%, 30%, and 76% for the other stock. Clearly, in both stocks, effects in mated females, when both female and male germ cells were treated, are relatively much higher than the added effects on dyctyate oocytes and spermatozoa. This is also true for the 25 mg/kg dose.


Induction of gene mutations and chromosomal aberrations by simian virus 40 in cultured mammalian cells.

Induction of gene mutations by SV40 was studied in aneuploid human and Chinese hamster cells. In Chinese hamster cells SV40-induced chromosome aberrations were also studied. SV40 penetrated into the cells of both lines and induced synthesis of the T antigen. The efficiency of infection in Chinese hamster cells was tested additionally by their ability to form colonies in medium lacking the serum growth factor. The maximal number of cells with serum growth factor independence was observed on the first day after infection. When hamster cells had been maintained in "factor-free medium" for the first two passages after infection a sub-line was isolated, which synthesized T antigen 60 days after exposure to SV40. This was considered to be an indirect proof of the integration of viral genome into host chromosome. A significant increase in the frequency of chromosomal aberrations was detected in SV40-infected Chinese hamster cells. It was observed on the first and second days after treatment. The most numberous were the chromosome and chromatid breaks, which were distributed randomly  in 5 morphological groups according to the chromosome length. SV40-induced mutations of resistance to 8-AG and 6-MP in human and Chinese hamster cells respectively were detected, when cells were plated in selective medium one to five days after infection. Induction was detected in all the 4 experiments with human cells and in 9 out of 11 experiments with Chinese hamster cells. Induction was highly significant according to the Wilcoxon test (P greater than 0.99), when the results of all experiments carried out in human and Chinese hamster cells were summarized. Resistance was stable after prolonged cultivation of 13 isolated clones under non-selective conditions. It is suggested that viral genome integration, gene mutations and chromosomal aberrations may have common molecular mechanisms. The role of gene mutations in virus-induced carcinogenesis is discussed.


Molecular structures of cytidine-5'-diphosphate and cytidine-5'-diphospho-choline, and their role in intermidiary metabolism.

The nucleotide coenzyme cytidine-5'-diphospho-choline is highly folded. The CMP-5' parts of the molecules in the crystal structure are strongly linked by metal ligation and hydrogen bonds leaving the phosphoryl-choline residues relatively free. Cytidine-5'-diphosphoric acid exists as a zwitterion with N31 protonated. The P-O bond lengths from the anhydride bridging oxygen in the pyrophosphate are significantly different.


Lipoprotein receptors and genetic control of cholesterol metabolism in cultured human cells.

Biochemical analysis of cultured fibroblasts obtained from human subjects with the autosomal dominant disorder familial hypercholesterolemia has led to the identification of a hitherto unrecognized cell-surface receptor for serum lipoproteins. This receptor plays a critical role in the physiologic regulation of cholesterol metabolism in human cells, and its absence leads to elevation of serum-cholesterol levels and severe atheroslerosis.


Cyclic AMP-dependent protein kinases and binding sites for cyclic AMP in rat erythrocytes.

In red cell preparations from reticulocyte-poor (untreated animals; approximately 2% reticulocytes) and reticulocyte-rich blood (animals pretreated with acetylphenylhydrazide; approximately 60% reticulocytes) of rats, cAMP binding sites and cAMP-dependent protein kinase activities were determined. High affinity binding sites for cAMP were present both in membrane and cytoplasmic preparations; while the apparent binding constants determined in both cell fractions (approximately 3 x 10(-9) M for membrane, approximately 2 x 10(-8) M for cytoplasmic fractions) were independent of the reticulocyte content of the preparations, the respective numbers of sites were about twice as high in the reticulocyte-rich as in the reticulocyte-poor preparations. In membrane preparations, significant cAMP-dependent protein kinase activity could be detected only in membrane fractions from reticulocyte-rich blood which were considerably contaminated by intracellular components ("haemoglobin-containing membranes') while in washed ("haemoglobin-free') membranes no cAMP-dependent protein kinase activity was found. In cytoplasmic preparations both from reticulocyte-poor and reticulocyte-rich blood, two different protein kinases, a low and a high Ka enzyme, were tentatively differentiated by kinetic data; the apparent activation constant for the high Ka enzyme (approximately less than 5 x 10(-8) M) was in the concentration range of the binding constants determined on cytoplasmic preparations. The activity of the high Ka protein kinase was several fold higher in reticulocyte-rich than in reticulocyte-poor cytoplasmic fractions, while the activity of the low Ka enzyme was obviously independent of the reticulocyte content. From the results obtained, it is concluded that in premature rat erythrocytes, membrane protein(s) may serve as protein substrates for cAMP-dependent protein kinase(s) located in the cytoplasm. This assumption was supported by experiments with intact erythrocytes (prelabelled with inorganic 32P-phosphate) from reticulocyte-rich blood: isoprenaline, theophylline, and also dibutyryl-cAMP significantly increased phosphorylation of membrane protein of these cells. From the results presented (and others previously reported) it becomes evident that only premature rat erythrocytes, i.e. reticulocytes, are equipped with a beta-adrenergic receptor-effector system consisting of a beta-adrenergically stimulated adenyl cyclase and cAMP-dependent protein kinase(s). Obviously, the adrenergic receptor system and also part of the effector system is lost during the process of red cell maturation.


cAMP as a second messenger in central blood pressure control.

Injections of dibutyryl cyclic adenosine 3',5'-monophosphate in doses of 125, 250 and 500 mug/kg into the lateral cerebral ventricle of anaesthetised cats produced dose dependent increases in blood pressure and heart rate, while doses of 50, 100 and 200 mug/kg imidazole-4-acetic acid, a stimulator of phosphodiesterase, exerted an opposite effect. The results are discussed in the light of the hypothesis that the activity of cardiovascular centres is determined by their concentration of cyclic AMP.


Characterisation of the adrenoceptor mediating changes in cyclic adenosine 3'-5' monophosphate in chick cerebral hemispheres.

Noradrenaline, adrenaline, isoprenaline and salbutamol induced marked accumulations of cyclic AMP in incubated slices of chick cerebral hemispheres. Isoprenaline was both more potent and more powerful than adrenaline or noradrenaline and the increase in cyclic AMP elicited by the catecholamines was powerfully antagonized by the beta-adrenoceptor antagonist propranolol but not by the alphapadrenoceptor blocker phentolamine. The order of potency of the catecholamines isoprenaline greater than adrenaline greater than noradrenaline and the ability of the non-catechol salbutamol to stimulate cyclic AMP accumulation in chick cerebral slices suggests that the beta-adrenoceptro may resemble that found in the lung (beta2) rather than that in the heart (beta1). Propranolol proved to be a very potent antagonist of the cyclic AMP response induced by isoprenaline in vivo. Beta-adrenoceptor blockade was still evident 12 hrs after a single injection of the durg although only negligible amounts of 3H-propranolol could be detected in cerebral tissue after 2 hrs.


Effects of drugs and axotomy on acetylcholine levels in central cholinergic neurons.

Administration of cholinergic nuscarinic agonists either did not alter, or increased hippocampal Ach levels, while antagonists lowered the levels. As observed previously, placement of lesions in the medial septal area resulted in a rise in hippocampal Ach levels in the first thirty minutes. Agonist drugs administered immediately before lesion, further increased the post-lesion rise in Ach levels. With antagonists, inconsistent results were obtained in that atropine blocked the post-lesion rise, while scopolamine and quinuclidinyl benzilate (QNB) potentiated the post-lesion rise. These results suggest that there may be receptor-mediated effects on Ach synthesis or release after axotomy.


Reactions of 4-halogeno-N,N-dimethylaniline-N-oxide with hemoglobin.

Incubation with ferrihemoglobin or ferricytochrome c transforms 4-bromo-N,N-dimethylaniline-N-oxide (BrDANO) into 4-bromo-N,N-dimethylaniline, 4-bromo-N-methylaniline, and 2-dimethylamino-5-bromo-phenol. In a secondary reaction 4-bromo-N'-(4-bromophenyl)-6-hydroxy-N,N,N-trimethyl-1,3-phenylenediamine (U) was found from 2-dimethylamino-5-bromo-phenol and 4-bromo-N-methylaniline. The products were isolated and identified, the structure of (U) was elucidated. It has been found that the formation of ferrihemoglobin by BrDANO is an autocatalytic reaction, ferrihemoglobin being the catalyst. BrDANO reacts with the catalyst resulting in the named products. 2-Dimetnylamino-5-bromo-phenol oxidizes hemoglobin in the presence of oxygen. Thereby the catalyst ferrihemoglobin increases, which in turn increases the transformation of BrDANO to ferrihemoglobin forming derivates. This cycyle explains the autocatalytic character of the production of ferrihemoglobulin by BrDANO. The primary products of the reaction between 4-fluoro-N,N-dimethylaniline-N-oxide (FDANO) or 4-chloro-N,N-dimethylaniline-N-oxide (ClDANO) with cytochrome c, 4-fluoro- and 4-chloro-N,N-dimethylaniline, 4-fluoro- and 4-chloro-N-methylaniline, 2-dimethylamino-5-fluoro-phenol, and 2-dimethylamino-5-chloro-phenol were isolated and identified. The reactions between N,N-dimethylaniline-N-oxide (DANO), BrDANO, ClDANO, FDANO, and hemoglobin from blood of various species were studied. BrDano and ClDANO accelerate the formation of ferrihemoglobin in solutions of hemoglobin from beef and dog blood much more effectively than DANO and FDANO.


Adrenal cortex adenylate cyclase: solubilization of adenylate cyclase and guanyl nucleotide binding sites.

Adenylate cyclase activity and binding sites for 5'-guanylyl-imidiphosphate [Cpp(NH)p] have been solubilized from bovine adrenal cortex membranes with Triton N-101. Solubilized adenylate cyclase is stimulated by Gpp(nh)p and fluoride ion but not by ACTH. Pretreatment of particulate membrane fractions with Gpp(NH)p, hormone and Gpp(NH)p or fluoride prior to detergent extraction leads to an activated solubilized enzyme which is nearly independent from added stimulants. Adenylate cyclase solubilized from membranes pretreated with Gpp(NH)p can be separated from guanyl nucleotide binding sites without loss of activity.


Effect of depolarizing agents on the adenosine-3',5'-monophosphate content of the bovine superior cervical ganglion.

The cAMP content of the bovine superior cervical ganglion increases rapidly when the tissue is incubated in media containing high concentrations of potassium and theophylline. This effect of high K+ is mimicked by Rb+, Cs+ and NH4+. The depolarizing substances veratridine and ouabain also cause an increase in the concentration of ganglionic cAMP; the effect of ouabain develops more slowly than that of veratridine. Tetracaine, in high concentrations, reduces the response to potassium, whereas it reduces the response to veratridine at much lower concentrations. Tetrodotoxin, at a concentration that barely affects the response to potassium, prevents the response to veratridine.


DNA content of mouse hepatoma nuclei determined by Feulgen cytophotometry.

The DNA content of mouse hepatoma nuclei was determined by means of Feulgen cytophotometry. The tumor nuclei showed a wide range of polyploidy and aneuploidy characteristic of malignant cells.


Scotomatous defects in the central visual field in pituitary adenomas.

In a series of 46 cases of pituitary adenomata (34 solid, 6 with yellow cyst, and 6 with hemorrhagic cyst), we found central scotomatous field defects in 8 patients. Of these 8 scotomatous cases 4 were associated with tumour containing a hemorrhagic cyst. We suggest that rapid expansion of the tumour is one of the principal etiological factors in the production of scotomatous instead of bitemporal field defects in patients with pituitary adenomas.


[Studies on cyclic 3', 5'-AMP system in human brain and its clinical application in Neurosurgical practice (author's transl)].

Literatures showed that cyclic AMP of cultured neoplastic cells of any kind was very low in concentration and also the effect of cyclic AMP and its derivatives on the malignant cells, especially on the malignant glioma, was already reported in vivo or in vitro from several neurosurgical units. The intrinsic content of cyclic AMP of the human cerebrum and the human brain tumors was first reported by authors in 1971. In this presentation the authors intended to confirm that the lower concentration of the cyclic AMP the more histologically malignant cerebral neoplasm, as well as in the cerebrospinal fluid, was observed. Concentration of cyclic AMP in the subcortical white matter, glioma, meningioma and medullobalstoma was much lower than in the gray matter tissue, however, it was not clear that the difference of the cyclic AMP concentration be possibly related to the malignancy of the human brain tumor. Furthermore, the cyclic AMP content of the cerebrospinal fluid of the patients with various brain tumor was not clearly different. The activity of adenyl cyclase was reported the highest in the synaptosome-containing fraction of the rat brain homogenate and this fact was significantly consistent with the finding that the highest concentration of the cyclic AMP was found in the human grey matter tissue. With the human brain gray matter authors determined successfully the activity of the human cerebral phosphodiesterase, which was probably localized in the post-synaptic membrane and was 158 nmole/mg protein/min. Its apparent Km was 0.9 x 10(-4) M. The results reported above have suggested the important participation of the cyclic AMP to cerebral synaptic transmission of nerve impulses, which was studied by light and electron-microscopic autoradiography utilizing the pulse labeling method with 3H-adenine. According to our study the majority of the adenyl cyclase of the human cerebrum was located synaptic structure and the finding obtained was quite compatible, as the first morphological study, with previously reported biochemical analyses. It was indicated that the cyclic AMP in the human brain was concerned to the cerebral synaptic transmission of nerve impulses and this should be very interesting and important to the clinical application for recovering cerebral function of neurosurgical patients.


[Viruses and brain tumors (author's transl)].

This brief review paper deals mainly with oncogenic DNA viruses originally isolated from human patients, i.e., human adenoviruses and human papova JC viruses. Human adenovirus type 12 was first isolated in 1953 in cell cultures derived from the adenoids of infected children. Since then, 32 antigenic types of human adenovirus have been identified. At least eight serotypes (12, 18, 31, 3, 7, 14, 16, and 21) are now known to be capable of producing tumors in newborn rodents. A direct causal relationship between a human adenovirus and malignant transformations in target cells (sensory neuronal precursors) has been definitely established by the development of a medullo-epitheliomatous neoplasm in the brain and spinal cord of an outbred strain of CD rats at as high an incidence as 90%. Intraocular inoculation of adenovirus in newborn rats within one week also has produced typical retinoblastomatous neoplasms. The remarkably uniform histopathologic appearance of all these malignancies in nervous tissue can be attributed to a primitive neuro-epitheliomatous neoplasm derived from sensory microneuron precursors that densely populate both the ventricular zone and the premature sensory retina at the point of virus inoculation. All of these brain and retinal tumors appear to share a common tumor phenotype, as all tumor cells contain cilia with the same morphology (a 9+0 pattern of doublets associated with a pair of centrioles). The production of adenovirus tumor-specific neoantigen (T), an earmark of the viral genome, can be regularly demonstrated by the immunofluorescein microscopic procedure. All transformed cells within both the ventricular zone and the retinal ganglion cell anlage thus appear to continue the production of (T) antigens. These findings lead us to assume that the target cell determinants in adenovirus tumorigenesis may reside in differentiating microneuron precursors ordained for the sensory neuronal complex.


Normal activity of metabolic pathways involved in the formation and utilization of phosphoribosylpyrophosphate in erythrocytes of patients with primary metabolic gout.

The activity of metabolic pathways involved in the formation and utilization of phosphoribosylpyrophosphate (PRPP) was studied in. The erythrocytes of 34 patients with idiopathic metabolic gout. The activities of the oxidative pentose shunt, of the hypoxanthine-guanine and adenine phosphoribosyltransferases (HGPRT, APRT) and of PRPP synthetase, as well as the rates of PRPP generation and of adenine incorporation into nucleotides were found to be normal in the erythrocytes of all these patients. Four patients with metabolic gout due to enzymatic abnormalities, two relatives with partial deficiency of HGPRT and two relatives with mutant feedback-resistant PRPP synthetase, were studied for comparison. The significance of the results is discussed in relation to postulated mechanisms for purine overproduction in metabolic gout.


[Peripheral nerve damage in upper extremities in glass cutters].

A group of glass cutters working with flint-glass was analysed in view of their susceptibility to ulnar nerve crush injury. The group comprised 429 cutters from Eastern Bohemia working in the Bohemia manufacture. The type of work done by cutters is described with particular reference to the way in which they rest their upper extremities during work and to the duration of work. Subjective symptoms were present in 44.7% of workers. The following neurological abnormalities were found during examination: hypaesthesia in 36.8%, motor activity impairment in 21.9%, muscular atrophy in 8.1%. Autonomic abnormalities involved hands in 29.1% of workers. Slight electromyographic abnormalities were demonstrated not only in the innervation area of the ulnar nerve but also in that of the median nerve. The authors discuss their views on the aetiology of the disease and criteria used in medical evaluation of ability to work in these workers.


Fetal lung maturatio. III. Amniotic fluid phosphatidic acid phosphohydrolase (PAPase) and its relation to the lecithin/sphingomyelin ratio.

The purpose of this study was to establish 1) the sequential changes in specific activity of phosphatidic acid phosphohydrolase (PAPase) in amniotic fluid and its relation to the lecithin/sphingomyelin (L/S) ratio; and 2) the origin of amniotic fluid PAPase. the increase in the amniotic fluid L/S ratio is preceded by an increase in PAPase activity, rising from 15 nmoles of phosphate released per milligram of protein per hour at 30 weeks to 100 nmoles at 37 weeks. The mean PAPase activity in the nasopharyngeal fluid of the infant is 456 nmoles of phosphate released per ml per hour, the amniotic fluid mean PAPase activity at delivery being 129 nmoles (P less than 0.01). These findings are consistent with the view that amniotic fluid PAPase originates, in part, from the fetal lung and likely participates in the regulation of the synthesis of lecithin.


The partial purification of Clostridium perfringens beta toxin.

An attempt was made to purify Clostridium perfringens Beta toxin. Crude toxin prepared by ammonium sulphate precipitation of culture supernatants was purified by chromatography on Sephadex G50, Sephadex G100 and DEAE cellulose. This material, although highly purified was not homogeneous on polyacrylamide gel electrophoresis. It had a toxicity of 800 000 mouse MLDs/mg N, a typical protein absorption spectrum in the UV region, an iso-electric point of 5, 6 and the main component had a molecular mass of 42 000 +/- 2 000 (estimated by electrophoresis in sodium dodecyl sulphate containing polyacrylamide gels).


[On a case of purulent pericarditis in a young child caused by mycobacterium rhodochrous].

M. rhodochrous was cultivated from the blood of a 2 1/2 year-old child with purulent pericarditis and is regarded as the causative agent. The process was successfully treated by chemotherapy and surgical measures. The problems of bacteriological diagnostics are discussed in detail with particular reference to the taxonomical differentiation of these agents and the treatment of their infections.


[Distribution of some enzymes in totally stained preparations of cestodes].

The distribution of succinic dehydrogenase, (see article), glucose-6-phosphat-dehydrogenase, alkaline and acid phosphatases and glucose-6-phosphatase was studied by means of the incubation of whole cestodes. Succinic dehydrogenase, NAD-diaphorase and glucose-6-phosphatdehydrogenase are connected in general with the fixating apparatus of the scolex and genital organs; phosphatases -- with the integument tissues, excretory system and calcareous corpuscles. The results obtained are in complete agreement with the available data on the distribution of the enzymes studied. The incubation method of whole cestodes can be useful for field works.


[Electron microscopic study of the excretory system of the hungry female tick Hyalomma asiaticum P. Sch. et E. Schl. Report II].

In H. asiaticum the cells of the Malpighian tubules and these of the rectal cas have the uniform structure: the apical surface is covered with microvilli, the basal plasmatic membrane forms relatively small invaginations. As to ultrastructural characters, there is no distinct division of the Malpighian tubule into departments. The distal ends of the tubules are only somewhat enlarged and form the so-called ampulla the cells of which are noticeably flattened. The microvilli and basal folds of the plasmatic membrane in this area of the tubule are indistinct. The cells of the ampulla and the neighbouring area of the tubule are characterized by the presence of inclusions with mucopolysaccharide secretion confined by the membrane. The microvilli are most developed on cells of the proximal ends of the Malpighian tubules. Well developed microvilli of the rectal sac form a striated border each containing a microtube inside. The basal invaginations are developed here better than in the cells of the Malpighian tubules.


[Short-term biological effects of synthetic salmon calcitonin in Paget's disease. Influence of posology].

In ten patients with Paget's disease of bone (Group I) intramuscular injection of 50 MRC units of synthetic Salmon calcitonin (SCT) induced a marked decrease of serum calcium (-1,444 mg%), serum phosphorus (-1,06 mg%), urinary total hydroxyproline (-71%), and a marked increase of urinary cyclic AMP (+ 114%). These changes occur at maximum 6 hours after the injection with a return to the initial values after 24 hours. In six other patients with Paget's disease (Group II) the acute biological effects were of the same nature and magnitude after the injection of 100 MRC units of SCT. In this group a significant but temporary increase of the plasma parathyroid hormone level was demonstrated. The magnitude of the hypocalcemia seems proportional to the initial serum alkaline phosphatase and urinary hydroxyproline levels. After one month of treatment, the alkaline phosphatase and urinary hydroxyproline have more marked and more regular decrease in group II who received 100 MRC units of SCT daily than in group I who received 50 MRC units three times a week. The cellular mechanisms of these biological changes are discussed. A posology of two daily injections of 50 MRC units of SCT is suggested when a quick and maximum stoppage of pagetic remodeling is required.


[Calcium and magnesium concentrations of the renal cortex in the rat. Comparison of 3 methods of tissular extraction of the principal cations].

We describe three methods for extraction of main tissular cations : a) dry ashing method; b) wet ashing method with HNO3; c) wet method on fresh tissue with HCIO4. This study allows the determination of tissular concentrations of calcium, magnesium in renal cortex of the Rat. Our findings show : a loss of Na, K cations probably by volatilization with dry ashing method ; inability for K determination when HCIO4 method is used ; tissular concentrations are : 4,7 mEq Ca ; 21,6 mEq Mg per kg of wet tissue (wet ashing method with HNO3) ; recovery percentage for magnesium is above 100 % (as previously shown by other authors) ; the dry ashing method is less suitable for extraction than the two other methods ; both wet methods lead to similar results and to a recovery percentage near 100 %.


[Herpes simplex virus antibodies in 18 patients submitted to immunosuppressive agents after renal transplantation].

The titer of circulating antibodies to herpes simplex virus type 1 was tested in 18 patients for a few months before and after renal transplantation. No clinical illness was observed in the group of patients lacking antibodies. On the contrary, half of those with antibodies developed cutaneous eruption within 2 to 4 weeks following treatment with immunosuppressive drugs. No relationship could be found between the appearance of clinical illness and the rise in antibody titer. This lack of correlation could be explained by the multiple modifications occurring in the immune system of the host submitted to immunosuppressive drugs.


Application of pulse polarography with anodic stripping voltammetry to biological and toxicological analyses for lead and cadmium.

The development of polarographic techniques within the last thirty years has made possible exceptional increases in sensitivity ; thus the order of concentration studied has passed from 10-3 g ion/1. to 10-10 g ion/1. Pulse polarography with anodic stripping voltammetry is, therefore, particularly suitable for the determination of a large number of the so-called " biologically  essential " and " highly toxic " metals, more often than not present in extremely low concentrations in biological samples which may be of limited volume (blood, for example). Our principal concern has been to describe and apply a method for the determination of lead and cadmium, sufficiently sensitive, precise, practical and rapid for everyday use by biologists and toxicologists. A fortiori this method is applicable in hydrology to the determination of traces of these metals in water.


[Mathematical study of size distribution of alveolar mononuclear cells (CMA) in man].

The mathematical study using the Wicksell transformation of the size distribution of the mononuclear alveolar cells has shown two different populations among the alveolar macrophages obtained from 4 adult men. These two types of cells could be also found out and described by electron microscopy. The large cells (14 microns in diameter) looked like the "alveolar macrophage ", with many phagolysosomes. The small cells (7 microns in diameter), less numerous (1/3 of the whole population) resembled monocytes. These data in man are compared to those found in rabbits and the role ot two kinds of cells is discussed.


[Epithelioid cell. Ultrastructural data during various pathological conditions].

A morphological study permitted us to show that the epithelioid cell is an active cell, the characteristic vesicles of which seem to be linked to the presence of a glycoprotein substance which passes through the Golgi apparatus and becomes accumlated in the lysosomes owing to their richness in acid phosphatase. A fraction of this substance may be secreted into the extra-cellular medium, without it being possible to determine its precise nature and whether this function is carried out only by this cell.


Fibroxanthosarcoma of bone.

A tumour arising from the body of the eighth thoracic vertebra is believed to be the first recordedexample of a primary fibroxanthosarcoma of bone. Apart from the absence of a demonstrable storiform pattern, the histology conformed to that of fibroxanthosarcoma of the soft tissues. Ultrastructural studies showed that asteroid-like bodies seen in the cytoplasm of some tumour cells consisted of aggregated centrioles surrounded by cytoplasmic vacuoles. The finding of Langerhans cell granules in some tumour cells supported their identification as histiocytes. These granules have not previously been recorded in the fibrous histiocytoma group of tumours.


Sphingomyelinases in human tissues. II. Absence of a specific enzyme from liver and brain of Niemann-Pick disease, type C.

Sphingomyelinase was obtained in excellent yield from liver and brain by homogenization with 0.05 M citrate-phosphate buffer, pH 4.5, containing 0.25% Triton-X-100 (v/v) followed by dialysis of the supernatant fluids against 1% glycine. Total recovery of enzyme was slightly less with tissue from Niemann-Pick disease compared with control tissue. Isoelectric focusing of liver and brain extracts was successfully used to resolve several species of sphingomyelinase. Three (I-III) of the five species were partially characterized. Enzyme I (pI 4.6) had a pH optimum of 4.8-5.0 in acetate buffer and a Km value of 0.026 mM. Both sphingomyelinases I and II were the major enzymes, whereas III, IV, and V were found at lower levels. Of the two major species in normal liver and brain (I and II), species I alone persisted in liver from the two cases of type C, while species III, IV, and V were present. In brain, only species II was decreased but the resolution of the brain enzymes was less satisfactory.


Sphingomyelinases in human tissues. III. Expression of Niemann-Pick disease in cultured skin fibroblasts.

Sphingomyelinase was effectively extracted cultured fibroblasts homogenized in 1% glycine. The average specific activity was 24.6 nmol substrate hydrolyzed per hr per mg protein. Cultured cells from two cases of Niemann-Pick disease type A and one case of type B had markedly reduced enzyme activity, whereas, in type E cells, total activity was twice normal. Sphingomyelinase was resolved by isoelectric focusing into three peaks of activity (I-II), where I and II were the major forms. Species I had a pI of 4.6, II a pI of 4.8, and III a pI of 5.0. All three peaks, although markedly reduced, were present in cell extracts from both types A and B. All species of enzyme in B cells were found at higher levels than in A cells. Cultured cells from a case of type E contained sphingomyelinase I at a level several fold higher than normal, species II was absent, whereas III was found at near normal levels.


Pyruvate dehydrogenase phosphatase deficiency: a cause of congenital chronic lactic acidosis in infancy.

A male child presented on the first day of life with metabolic acidosis with elevated blood lactate (15 mM), pyruvate (0.4 mM), and free fatty acid (1.3 mM) levels and a blood pH of 7.16. The severity of the acidosis was diminished by intravenous administration of glucose in large doses and by bicarbonate. On two occasions, when the acidosis was particularly severe, peritoneal dialysis using an acetate buffer was required. Restriction of the dietary intake of saturated fatty acids or treatment with nicotinic acid also appeared to diminish the severity of acidosis. No improvement was achieved by the administration of thiamine or biotin. Tissues taken at postmortem showed normal activity of gluconeogenic enzymes and pyruvate dehydrogenase. The activity of pyruvate dehydrogenase in tissue homogenates preincubated with ATP was reduced by 60-75% both in liver of the patient and of the controls because of the inactivation of the enzyme by pyruvate dehydrogenase kinase. Addition of Ca++ and Mg++ to the inactivated enzyme caused a prompt return of the activity to normal in controls but not in the patient. This defect, which was apparent in muscle and liver but not in brain, we attribute to a markedly reduced activity of pyruvate dehydrogenase phosphatase in the patient.


Demonstration of human leukocyte degranulation induced by sera from homozygotes and heterozygotes for cystic fibrosis.

The ability of epsilon-amino caproic acid (EACA)-treated normal serum and of cystic fibrosis (CF)-affected and carrier sera to promote the release of lysosomal enzymes from sensitized human polymorphonuclear leukocytes (PMN) was assessed through the measurement of beta-glucuronidase and myeloperoxidase activity after exposure of these cells to the various test sera. This study was initiated to extend the analogies between preciliary dyskinesia factor (pre-CDF), separated from the cell-free media of cultures derived from CF homozygous and heterozygous individuals, and C3a anaphylatoxin. The extent of lysosomal degranulation of human PMN exposed to fresh untreated sera of each of five controls, seven CF homozygotes, and eight heterozygotes, as expressed by the amount of beta-glucuronidase releases, was 7.84% (+/- 0.934) for countrol sera, 14.01% (+/- 1.79) for CF-affected sera, and 10.61% (+/- 1.43) for heterozygous sera. The difference between CF homozygotes and control subjects is significatn (P less than 0.0001), as is the difference between CF-affected and carrier individuals (0.001 less than P less than 0.005) and between control subjects and carriers (0.001 less than P less than 0.005), when beta-glucuronidase. However, the differences between control subjects and CF heterozygous individuals are not significant. Treatment of these sera with 1 M EACA gave values for beta-glucuronidase and myeloperoxidase release which are slightly reduced when compared with those obtained with fresh, untreated samples. EACA apparently reduces the activity of beta-glucuronidase released from PMN. Amicon filtration studies of these serum samples demonstrated that degranulating ability and the presence of cilicary dyskinesia, as assessed by rabbit tracheal bioassay, are not always associated. Therefore, the relationship between pre-CDF and the degranulator activity in native CF-affected and carrier sera is unclear, in part because of the limitations inherent in the test systems employed.


Seroepidemiologic survey of varicella: Value of specific fluorescent antibody test.

The sensitive varicella-zoster fluorescent antibody to membrane antigen (V-Z FAMA) test was used in a seroepidemiologic survey of two outbreaks of varicella involving 30 children. The attack rate of varicella based on clinical observations alone was 60%. The attack rate based on clinical and serological observations lay between 78% and 82%, after excluding those children who had detectable antibody at the time of exposure. No subclinical varicella infection was observed in this small series of patients. No boost in antibody response was observed in immunes who were exposed to varicella. No patients with detectable V-Z antibody by FAMA at the time of exposure developed clinical varicella. The V-Z FAMA test provided helpful information about the immune status of persons exposed to varicella.


A comparison of the physical and intellectual development of black children with and without sickle-cell trait.

Sickle-cell trait, a condition present in 7% to 9% of the United States Black population, is usually considered to be a clinically benign condition. However, there is increasing evidence to indicate the contrary, that is, the clinical pathophysiology is variable, ranging from a benign condition in most cases to a relatively few cases of severe pathological involvement. Physical and intellectual growth measures were taken on 19 children with sickle-cell trait (12 boys and 7 girls) from a large study of Black same-sex twin pairs from Philadelphia, Pennsylvania, and compared to measures taken of a sample of normal Black children from 155 monozygotic and dizygotic same-sex twin pairs. Sickle-cell trait carriers were found to weigh less, have smaller upper arm circumference lesser skinfold thickness, and showed less mature skeletal age, differing significantly from normal children. Sickle-cell carriers tended to score lower on four of five intellectual measures, scoring one fifth to one third of a standard deviation lower than normal children.


The RWT method for the prediction of adult stature.

The Roche- Wainer- Thissen (RWT) method estimates the adult stature of an individual from data recorded at a single childhood examination. The data required are recumbent length, nude weight, midparent stature, and hand-wrist skeletal age. If necessary, a measurement of the child's stature can be transformed to be approximately equivalent to recumbent length with little loss of accuracy. When applied to data from three longitudinal growth studies, the prediction errors with the RWT method were smaller than those with the method of Bayley and Pinneau.


Cyclic AMP and the potassium-induced vasodilation.

The cAMP concentration of bovine facial arteries (helical strips) increased from 0.84 +/- 0.14 to 1.51 +/- 0.24 nmole/g wet weight, when the strips were relaxed by exposure to a physiological salt solution containing 10 instead of 2.7 mM KCl. A similar result (increase from 1.13 +/- 0.15 to 2.07 +/- 0.29 nmole cAMP/g wet weight) was obtained in strips pretreated with 10(-7) g/ml beta-methyldigoxin. These strips reacted to 10 mM KCl with a constriction. Addition of the phosphodiesterase activator N-methyl-imidazole (3 - 10(-3)M) did not affect the mechanical response to 10 mM KCl. It is concluded that for the potassium-induced vasodilation cAMP does not have a mediating function comparable to that in beta-adrenergic relaxation of vascular smooth muscle.


Contribution of long chain fatty acids to the energy supply of the rat kidney cortex.

Tubular fragments from rat kidney cortex were isolated by collagenase and suspended in an incubation medium containing a combination of several renal substrates. Substrate concentrations were in the physiological range. O2 uptake, total CO2 production, and the 14CO2 production from U-14C-labeled palmitate and oleate were measured. During the first minutes of incubation the CO2 production from palmitate and oleate was 10.5% or 6.3%, respectively, of the total CO3 produced. The RQ was 0.897. A subsequent decrease of the total CO2 production at a constant uptake of oxygen indicated a rising contribution of fatty acids to the fuel of respiration. The renal preference for substrates other than longchain fatty acids is discussed.


Vasopressin dependent adenylate cyclase in single segments of rabbit kidney tubule.

AVP dependent adenylate cyclase activity was measured in single pieces of 8 different tubular segments isolated from collagenase treated rabbit kidneys. High responses were observed in all the tested portions of the collecting tubule, that is its cortical branched part (BCT), its cortical straight part (CCT) and its outer medullary part (MCT). Dose response curves indicated in CCT: 2 fold threshold stimulation at 10(-11) M AVP, 27 fold stimulation at 10(-6) M AVP, half maximal stimulation at about 10(-9) M AVP. Both the medullary (MAL) and, to a lesser extent, the cortical (CAL) portions of the thick ascending limb were also observed to contain AVP sensitive adenylate cyclase (for MAL: 2 fold threshold stimulation at 10(-9) M AVP, 9 fold stimulation at 10(-7) M AVP, half maximal stimulation at 5 X 10(-9) M AVP). In contrast, nearly no responsiveness to AVP was observed in the proximal convoluted tubule, in the thin descending limb of the loop and in the distal convoluted tubule (DCT). The limited response obtained in DCT (which is a structure generally considered as a target site for AVP) as well as the clearcut effect elicited by AVP in MAL (the functioning of which is not known to be controlled by ADH) were expected observations; their possible physiological implications will be discussed.