Alternate pathway activation of complement in a Proteus mirabilis ulceration of the cornea. A 63-year-old patient had an extensive infiltration, ulceration, and eventual perforation of the cornea caused by Proteus mirabilis. Histopathological examination of corneal tissue that was obtained at keratoplasty disclosed that the stroma was ulcerated with a diffuse infiltration of polymorphonuclear leukocytes. Immunopathological examination of the cornea revealed prominent and diffuse staining for properdin and C3 complement. There was an absence of staining for IgG, IgA, IgM, IgE, and C4 complement. These findings suggest that the alternate pathway of complement was activated in this ulcerated cornea and that immunopathological phenomena contribute to the polymorphonuclear infiltration found with Gram-negative ulcerations of the human cornea.


Factors in the spread of salmonellas in meatworks with special reference to contamination of knives. Immersion of knives momentarily in hot water (82 degrees C) was ineffective in destroying salmonellas on knives used in a meatworks to carry out the bung dropping operation. Laboratory experiments confirmed that knives covered with meat products required 10 or more seconds to be effectively decontaminated at this temperature. Examination of knives used for slaughtering and for dressing beef carcases showed that knives coming into contact with hides had higher counts for salmonella and a higher percentage positive than knives used for other cutting operations. Knives used for cutting the skin of the forelegs and hindlegs had the highest counts.


Subdivision of Mycoplasma mycoides subsp. mycoides from cattle and goats into two types. Some strains of Mycoplasma mycoides subsp. mycoides, mostly isolated from goats, grow to greater turbidity in broth and form larger colonies on solid medium than does the type strain, PG1. These strains also digest casein, liquefy inspissated serum actively and survive longer at 45 degrees C and are referred to as LC (large colony) strains. Strains more closely resembling PG1 have been called SC (small colony) strains. The SC strains comprise those from contagious bovine pleuropneumonia (CBPP) and some from goats. One LC strain was isolated from a steer; all others have come from goats. Differentiation of M. mycoides subsp. mycoides into 2 types on the basis of the characteristics described may be relevant to their role in CBPP.


Binding of chloramphenicol and a fragment of aminoacyl-transfer ribonucleic acid to ribosomes and a ribosome precursor from a mutant of Escherichia coli. During exponential growth, the mutatn strain Escherichia coli 15-28 accumulates 47S particles, which are unusual precursors to 50S ribosomal subunits. The 47S particles have little ability to bind chloramphenicol, but binding of a fragment of aminoacyl-tRNA is about half that by completed subunits. The 70S (and 50S) ribosomes of strain 15-28 and its parent (strain 15TP) do not differ in chloramphenicol binding. Although ribosomes from the mutant are less able than those from the parent to bind the fragment, this difference is not as marked as was found previously [Sims & Wild (1976) Biochem. J. 160, 721-726] for the binding of an analogue of peptidyl-tRNA and for peptidyltransferase activity. The altered activities may arise because strain 15-28 misassembles 50S subunits of altered conformation and because the few proteins that 47S patricles lack have vital functions in some of the partial reactions of protein synthesis.


Proline biosynthesis by cell-free extracts of Escherichia coli and potential errors arising from the use of a bioradiological assay procedure. 1. The growth of Escherichia coli proline auxotrophs on medium containing L-proline (50 microgram/ml) induces catabolic enzymes. A bioradiological assay system for proline, using proB cells of E. coli, might give erroneous results owing to proline catabolism by the proline auxotrophs on which the assay depends. 2. Differential utilization of proline and 1-pyrroline-5-carboxylate by the proB cells for the synthesis of protein, and failure of the method to distinguish between these two possible products of the proline-biosynthetic enzymes, might also give rise to error. 3. The proline-dependent incorporation of [14C]phenylalanine into the protein of proline-starved proB auxotrophs was to some degree directly influenced by the presence of crude cell extract from E. coli, even though this was not supplied with substrate and cofactors, and could thus not itself synthesize proline. 4. The kinetics of proline biosynthesis by cell-free extracts were linear and biphasic, only the last phase being affected by the concentrations of substrate and extract. This phenomenon is not understood. 5. Proline biosynthesis is inhibited, not only by high concentrations of ATP, but also by aspartate, glycine, alanine and serine, aspartate having the greatest effect. 6. Attempts at complementation in vitro between extracts of proline auxotrophic mutants were not successful, suggesting the possibility that strain X680 (proA) and/or X278 (proB) may be a double mutant. 7. The enzymes of proline biosyntehsis are co-eluted from a column of Bio-Gel A1.5M in a position corresponding to a mol.wt. of 350000. 8. Comparisons between rates of proline biosynthesis in vivo and in vitro were made.


Gel filtration of sialoglycoproteins.

Gel filtration of sialoglycoproteins. The role of sialic acid in the gel-filtration behaviour of sialoglycoproteins was investigated by using the separated isoenzymes of purified human liver alpha-L-fucosidase and several other well-known sialic acid-containing glycoproteins (fetuin, alpha1-acid glycoprotein, thyroglobulin and bovine submaxillary mucin). For each glycoprotein studied, gel filtration of its desialylated derivative gave an apparent molecular weights much less than that expected just from removal of sialic acid. For the lower-molecular-weight glycoproteins (fetuin and alpha1-acid glyocprotein), gel filtration of the sialylated molecules led to apparent molecular weights much larger than the known values. The data indicate that gel filtration cannot be used for accurately determining the molecular weights of at least some sialoglycoproteins.


Solubilization of bovine heart-value collagen. Complete solubilization of bovine heart-valve gollagen was obtained by using sequential pepsin digestion and extraction with dithiothreitol under non-denaturing conditions. Use of the reducing agent was the crucial step resulting in solubilization. These findings suggest that disulphide-bonded proteins may be in part responsible for the insolubility of bovine heart-valve collagen.


[Composition of the 32-detector rCBF system (Meditronic) and experience in its use (author's transl)]. Regional cerebral blood flow (rCBF) was measured in patients with no neurological deficit and no abnormal findings of cerebral angiograms except aneurysm itself with Xenon-133 injection technique by using the computerized 32-detector rCBF system (rCBF-322, Meditronic, Denmark). This system consists of a 32-detector head with a concave surface and a 32 pulse height analyser and analog-digital ratemeter as the accumulator connected to display units with an oscilloscope for 32 clearance curves and also a microcomputer programmed for calculating the rCBF initial with the initial slope analysis, for calculating the rCBF 10 with the height-over-area method and the rCBF gray, rCBF white, weight gray and weight white with the SHAM method. The results obtained were as follows: 1) The mean value and SD of rCBF initial was 58.4 +/- 6.2 ml/100 g brain/min; rCBF 10, 50.6 +/- 5.0 ml/100 g brain/min; rCBF gray, 78.0 +/- 14.0 ml/100 g brain/min; rCBF white, 22.1 +/- 6.6 ml/100 g brain/min; weight gray, 49.0 +/- 3.7%; weight white, 51.0 +/- 3.7%. These data should be considered normal values as reported also by others. 2) Reproducibilities were estimated by measuring twice the rCBF initial and the rCBF 10 at rest with closed eyes. The reproducibilities of the rCBF initial eyes 13.55% (P less than 0.05) and one of the rCBF 10 was 8.85% (P less than 0.05). It was concluded that this system should be widely used for measuring the rCBF in patients with various cerebral diseases.


Hypertension and arterial development. Long-term considerations. A considerable body of evidence supports the view that haemodynamic factors, acting during growth and development, affect the ultimate form of the larger arteries. Many of the changes induced in vessels by high pressure or flow are adaptive and effectively minimise alterations in vessel function, but these changes may predispose to degenerative disease. 'Early intervention' in arterial disease implies intervention in the first and second decades of life.


Comparative trial of atenolol and propranolol in hyperthyroidism. 1 The use of atenolol, a cardioselective beta-adrenoceptor antagonist, in the management of hyperthyroidism has been studied by comparing it with propranolol. 2 In a double-blind cross-over trial, atenolol (50 mg), propranolol (40 mg) and placebo 4 times daily for 1 week were compared in twenty-one hyperthyroid patients by sequential analysis. 3. Patients generally preferred atenolol or propranolol to placebo but this preference only achieved significance with propranolol. 4 Judged by their effect on the symptoms and signs of hyperthyroidism, both atenolol and propranolol were significantly better than placebo, but no distinction could be made between the two active compounds. 5 Atenolol and propranolol reduced mean heart rate by 29.8 and 27.1% respectively compared with placebo. 6 Atenolol appeared almost equally effective to propranolol in the management of the peripheral manifestations of hyperthyroidism.


Phenotypic diversity in experimental hepatomas: the concept of partially blocked ontogeny. The 10th Walter Hubert Lecture. Cancer cells should be seen not as exclusively a problem in cell proliferation, but rather as a problem combining the processes of proliferation and differentiation, hence the phrase introduced in 1968: "oncogeny is blocked ontogeny". Cancer tissues resemble foetal tissues in many ways but they differ from foetal tissue in being unable to "recapitulate the total programme leading to an orchestrated collection of organism-serving cells" that are programmed "to make the organ as adaptive as possible to the range of environmental variations in which it evolved". Citing the "Osgood Principle" from the 1950's, recent supporting evidence was described, in which the most mature differentiated cells exert positive and negative feedback upon the proliferation of their progenitor stem cells. Advanced examples in the haemopoietic series were drawn from the work of Sachs, Metcalf, Till and McCulloch, and Kurland and Moore. The blocked ontogeny hypothesis was further elaborated in the concept of "partially-blocked ontogeny", which is intended to describe a situation in which highly differentiated slowly growing tumours contain some cells which have left the proliferating pool to differentiate along the normal pathway, but are blocked somewhere short of the final organism-serving state, in harmony with earlier suggestions by Osgood, by Pierce, and by Sachs.


Rapid serological test for diagnosis of chlamydial ocular infections. A rapid serodiagnostic test for the diagnosis of paratrachoma (TRIC ophthalmia neonatorum, inclusion conjunctivitis, TRIC punctate keratoconjunctivitis, and trachoma of sexually transmitted origin) has been developed. The technique is based on using a modified micro-immunofluorescence test for detecting antichlamydial IgG and IgM in the blood and IgG and IgA in tears. The blood samples are collected on cellulose sponges after a finger prick, and tears are collected by introducing small sponges into the lower conjunctival fornix of the eye. The blood and tear samples collected in this way could be sent to the diagnostic laboratory by post without special arrangements for cold storage. In general the presence of antichlamydial IgG at a level of greater than or equal to 1/32 or IgM at a level of greater than or equal to 1/8 in blood and antichlamydial IgG or IgA at a level of greater than or equal to 1/8 in tears was closely associated with ocular paratrachoma. The combined results of the micro-IF test of blood and tears has yielded the highest rate of positivity (90%). In patients with acute untreated paratrachoma the sensitivity of this test was similar to that of irradiated McCoy cells. In patients with a milder infection receiving antibiotics the sensitivity of the serodiagnostic test was superior to that of the cultural test. The high sensitivity and specificity of this rapid, simple and inexpensive serodiagnostic test for the diagnosis of chlamydial ocular infections, coupled with simple and practical methods of collection and transport of blood and tear specimens, offer advantages over cultural tests for routine diagnosis and study of chlamydial ocular infections.


Ocular histopathology in animals experimentally infected with Mycobacterium leprae and M. lepraemurium. 1. Mycobacterium leprae and M. lepraemurium infections in the mouse. 2. Mycobacterium leprae infections in the 9-banded armadillo (Dasypus novemcinctus L.). At varying periods of time following the successful establishment of systemic infections with Mycobacterium leprae or M. lepraemurium in the mouse and the nine-banded armadillo eyes were examined by light microscopy. Inoculation of bacilli was by the intravenous or intraperitoneal route or directly into the hind footpads; eyes were not directly inoculated in this study. During periods of up to 3 years under laboratory conditions no animal showed evidence of impaired vision or blindness, and the external appearance of both eyes was normal. The ocular histopathology and the sites of accumulation of bacilli are described. In immunologically normal mice infected with M. lepraemurium bacilli were much commoner in extraorbital tissues, but they were, nevertheless, found in various tissues within the orbit, including the ciliary body and sclera. In immunologically normal mice (and one rat) injected with M. leprae of human origin no bacilli were found in the eye, but in mice immunologically depressed by thymectomy and total body irradiation considerable numbers of bacilli were present in the iris and ciliary body and also in the limbal cornea. In the armadillo bacilli were found in large numbers in virtually all tissues except the lens, retina, optic nerve, and aqueous and vitreous humours, but the uveal tract was heavily involved. Findings are discussed in relation to the great frequency of ocular involvement and the importance of immune-complex disease in patients with lepromatous leprosy, and to factors wihch may favour the localisation and multiplication of Mycobacterium leprae in the eye.


Replacement of pseudouridine in transfer RNA by 5-fluorouridine does not affect the ability to stimulate the synthesis of guanosine 5'-triphosphate 3'-diphosphate. The requirement for ribothymidine and pseudouridine in the TpsiCG loop of tRNA for its activity in the ribosome and tRNA-stimulated synthesis of guanosine 5'-triphosphate 3'-diphosphate (pppGpp) by stringent factor has been tested by the use of a purified tRNAPhe (883 pmol of phenylalanine incorporated/A260 unit) in which 92% of the pseudouridine, 98% of the ribothymidine, 98% of the dihydrouridine, and 88% of the uridines were substituted by 5-fluorouridine. This tRNA was quantitatively as active as control tRNA in inducing pppGpp synthesis. With loose-couple ribosomes, the concentration of tRNA needed to give half-maximal reaction was 0.07 micrometer for both normal and fluorouridine-substituted tRNA, with vacant tight-couple ribosomes it was 0.05 micrometer, and with tight couples carrying poly(Phe)-tRNA at the P site the value was 0.02 micrometer. These results show that at the level of intact tRNA there is no special requirement for modified bases in the TpsiCG loop of tRNA in the synthesis of pppGpp.


Control of acetohydroxy acid synthetase in Escherichia coli 9723. A method by which three acetohydroxy acid synthetase activities are separated from extracts of Escherichia coli 9723 has been developed. Isoleucine specifically represses synthesis of one of the enzymes, which is not sensitive to valine inhibition, and isoleucine also simultaneously enhances the production of a second activity, which is valine inhibitable. The valine-inhibitable activity is repressed by leucine and valine, a combination of which is more effective than either alone. The third acetohydroxy acid synthetase, which is more active at pH 6 than at 8, is not controlled by the branched-chain amino acids. In a mutant of E. coli 9723 selected for the ability of valine to inhibit growth, the isoleucine-repressible acetohydroxy acid synthetase activity was no longer present, but isoleucine addition still resulted in enhanced production of the valine-inhibitable activity.


Photoaffinity inhibition of dipeptide transport in Escherichia coli. A dipeptide containing a nitrene precursor, glycyl-4-azido-2-nitro-L-phenylalanine, has been synthesized. This compound is a photoaffinity inhibitor of dipeptide transport in E. coli. In the dark, the dipeptide is a reversible inhibitor of glycylglycine uptake by live E. coli W cells. The 14C-labeled compound is a substrate for the transport system, with a Km of 7 micrometer and V max of 5 x 10(3) molecules cell-1 s-1 (compare 9 micrometer and 1 x 10(4) molecules cell-1 s-1, respectively, for the transport of glycylglycine under the same conditions). When intact E. coli cells are photolyzed at approximately  350 nm in the presence of the photolabile dipeptide, their ability to transport either glycylglycine or unphotolyzed glycyl-4-azido-2-nitro-L-phenylalanine is irreversibly inhibited, but their ability to transport arginine is unaffected. The presence of glycylglycine in the medium during photolysis protects the cells against the light-dependent inactivation of dipeptide transport.


A new method for the purification of 30S ribosomal proteins from Escherichia coli using nondenaturing conditions. A new method for the purification of Escherichia coli (A19) 30S ribosomal proteins has been developed that avoids the use of denaturing conditions such as urea, acetic acid, and lyophilization. In this way the majority of the proteins from the small ribosomal subunit can be obtained in 5--100 mg quantities and at greater than or equal to 90% purity. This has been achieved by the initial "splitting" of the proteins into two main groups with LiCl followed by fractionating on ion-exchange and gel-filtration columns, in the absence of urea and in the presence of salt. The proteins prepared by this nondenaturing procedure were soluble at high ionic strength and less soluble, being aggregated, at low salt concentrations. This behavior was exactly the opposite of that exhibited by proteins prepared with methods using denaturing conditions. These new methods have enabled additional ribosomal RNA-binding proteins to be found and potential protein-proteins complexes to be isolated. Preliminary evidence that these proteins may retain a more native structure is presented.


Tripeptide-specific aminopeptidase from Escherichia coli AJ005. A tripeptidase, TP, from the ribosome-free fraction of Escherichia coli AJ005, a peptidase-deficient mutant of strain K-12, has been obtained using gel electrophoresis and chromatography on DEAE-Sephadex A-50, hydroxylapatite, and Sephadex G-200. Characterization studies on tripeptidase TP, freed of other detectable peptidases, indicate that this enzyme is capable of cleaving an amino-terminal leucine, lysine, methionine, or phenylalanine residue from certain tripeptides. Only one band of activity toward several tripeptides (and no activity toward dipeptides) was detected following gel electrophoresis of this preparation. Tripeptidase TP, the only strain AJ005 peptidase known to attack trilysine, was inactive toward all dipeptides, peptide amides, substituted peptides, esters, and tetrapeptides tested as substrates. Trilysine cleavage is optimal at about pH 8.5,  as determined in Tris, borate, or phosphate buffers. Tripeptidase TP activity tested under a number of conditions was not inhibited by soybean trypsin inhibitor (3 mg/mL), phenylmethanesulfonyl fluoride (25 micrometer), or iodoacetate (9 mM). p-Mercuribenzoate (10 micrometer), divalent copper, cobalt, calcium (2.5 mM), zinc (25 micrometer), and mercury (10 micrometer) are inhibitory. Based on Sephadex G-200 chromatography tripeptidase TP has a particle weight of approximately 80 000 daltons. An apparent Km of 5.3 mM was determined for methionylglycylglycine cleavage.


Aminoacyl-tRNA synthetases from yeast: generality of chemical proofreading in the prevention of misaminoacylation of tRNA. The specificity of valyl-, phenylalanyl-, and tyrosyl-tRNA synthetases from yeast has been examined by a series of stringent tests designed to eliminate the possibility of artefactual interference. Valyl-tRNA synthetase, as well as activating a number of amino acid analogues, will accept alanine, cysteine, isoleucine, and serine in addition to threonine as substrates for both ATP-PPi exchange and transfer to some tRNAVal species. The transfer is not observed if atempts are made to isolate the appropriate aminoacyl-tRNAVal-C-C-A but its role in the overall aminoacylation can be suspected from both the formation of a stable aminoacyl-tRNAVal-C-C-A(3'NH2) compound and from the stoichiometry of ATP hydrolysis during the aminoacylation of the native tRNA. Similar tests with phenylalanyl-tRNA synthetase indicate that this enzyme will also activate and transfer other naturally occurring amino acids, namely, leucine, methionine, and tyrosine. The tyrosine enzyme, which lacks the hydrolytic capacity of the other two enzymes (von der Haar, F., & Cramer, F (1976) Biochemistry 15, 4131--4138) is probably absolutely specific for tyrosine. It is concluded that chemical proofreading, in terms of an enzymatic hydrolysis of a misacylated tRNA, plays an important part in maintaining the specificity in the overall reaction and that this activity may be more widespread than has so far been suspected.


Formation of large, ion-permeable membrane channels by the matrix protein (porin) of Escherichia coli. One of the major proteins of the outer membrane of Escherichia coli, the matrix protein (porin), has been isolated by detergent solubilisation. When the protein is added in concentrations of the order 10 ng/cm3 to the outer phases of a planar lipid bilayer membrane, the membrane conductance increases by many orders of magnitude. At lower protein concentrations the conductance increases in a stepwise fashion, the single conductance increment being about 2 nS (1 nS = 10(-9) siemens = 10(-9) omega -1) in 1 MKCl. The conductance pathway has an ohmic current vs. voltage character and a poor selectivity for chloride and the alkali ions. These findings are consistent with the assumption that the protein forms large aqueous channels in the membrane. From the average value of the single-channel conductance a channel diameter of about 0.9 nm is estimated. This channel size is consistent with the sugar permeability which has been reported for lipid vesicles reconstituted in the presence of the protein.


Purification and properties of trehalase from the thermophilic fungus Humicola lanuginosa. Trehalase (alpha,alpha-Trehalose glucohydrolase, EC 3.2.1.28) was partially solubilized from the thermophilic fungus Humicola lanuginosa RM-B, and purified 184-fold. The purified enzyme was optimally active at 50 degrees C in acetate buffer at pH 5.5. It was highly specific for alpha,alpha-trehalose and had an apparent Km = 0.4 mM at 50 degrees C. None of the other disaccharides tested either inhibited or activated the enzyme. The molecular weight of the enzyme was around 170 000. Trehalase from mycelium grown at 40 and 50 degrees C had similar properties. The purified enzyme, in contrast to that in the crude-cell free extract, was less stable. At low concentration, purified trehalase was afforded protection against heat-inactivation by "protection against heat-inactivation by "protective factor(s)" present in mycelial extracts. The "protective factor(s)" was sensitive to proteolytic digestion. It was not diffusible and was stable to boiling for at least 30 min. Bovine serum albumin and casein also protected the enzyme from heat-inactivation.


Selective denaturation of several yeast enzymes by free fatty acids. The denaturation of eight purified yeast enzymes, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, 3-phosphoglycerate kinase, alcohol dehydrogenase, beta-fructosidase, hexokinase and glucose-6-phosphate isomerase, promoted under controlled conditions by the free fatty acids myristic and oleic, is selective. Glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate:NADP+ 1 oxidoreductase, EC 1.1.1.49) is extremely sensitive to destabilization and was studied in greater detail. Results show that chain length and degree of unsaturation of fatty acids are important to their destabilizing effect, and that ligands of the enzyme can afford protection. The denaturation process results in more than one altered form. These results can be viewed in the perspective of the possibility that amphipathic substances, and in particular free fatty acids, may play a role for enzyme degradation in vivo, by initiating steps of selective denaturation.


Influence of viscosity of enzymatic reactions studied with glucoamylase of Aspergillus niger. Viscosity can be interpreted in terms of transport of momentum and, therefore, it should influence the kinetics of enzyme reactions. A theory, developed by Somogyi and Damjanovich ((1975) J. Theor. Biol. 51, 393--401), is based on this idea. Transport of momentum must always be accompanied by the transport of mass and this second influence of viscosity is a limiting factor for fast reactions in the liquid phase. A third aspect is, that the chemical potentials of the components of viscous solutions are altered. This paper reports experiments concerning the influence of the viscosigens (compounds that increase the viscosity of solvents), alginate, sucrose, and maltose on the kinetic behaviour of glucoamylase (1,4-alpha-D-glucan glucohydrolase, EC 3.2.1.3). The observed invariance of V and the decrease of Km are explained by the increase of chemical potentials and restriction of momentum transport.


Enhanced allosteric regulation of threonine deaminase and acetohydroxy acid synthase from Escherichia coli in a permeabilized-cell assay system. A permeabilized-cell technique for rapid assay of enzyme activity has revealed enhanced allosteric regulation of both threonine deaminase (L-threonine hydrolyase (deaminating), EC 4.2.1.16) and acethohydroxy acid synthease (acetolactate pyruvate-lyase (carboxylating), EC 4.1.3.18) in Escherichia col K-12. In the permeabilized cell assay threonine deaminase exhibited a higher Hill coefficient for inhibition by L-isoleucine, and acetohydroxy acid synthase exhibited a hypersensensitivity to allosteric inhibition by L-valine when compared to studies on crude extracts. We propose that these effects reflect the in situ microenvironments of both enzymes. Preliminary evidence further indicates that acetohydroxy acid synthase may loosely associate with the cell membrane.


Triacylglycerol synthesis in lipid particles from baker's yeast (Saccharomyces cerevisiae). Triacylglycerol synthesis has been studied in a lipid particle preparation of baker's yeast (Saccharomyces cerevisiae), and compared with the synthesis in other subcellular fractions. Fatty acid-CoA ligase (AMP) (EC 6.2.1.3) activity and sn-glycerol 3-phosphate acyltransferase activity (EC 2.3.1.15) were present in all the subcellular fractions tested but the highest specific activities of both enzymes were observed with the lipid particle fraction. The products of the glycerol 3-phosphate acylation indicate that triacyglycerol synthesis proceeds through the phosphatidic acid pathway. However, only a small and nearly constant amount of lysophosphatidic acid was found with the lipid particle fraction while the other subcellular fraction produced lysophosphatidic and phosphatidic acid with a more pronounced precursor/product relationship. Triacylglycerol synthesis from endogenous diacylglycerol present in the lipid particle was also demonstrated.


Ultrastructural localization of beta-D-galactan in the nuclei of the myxomycete Physarum polycephalum. The acellular slime mold Physarum polycephalum produces an extracellular sulfated and phosphorylated beta-D-galactan which was recently isolated from the nuclei of this organism. This polysaccharide has now been localized in the nuclei of P. polycephalum by electron microscopy using a specific "sandwich" technique: thin sections of P. polycephalum microplasmodia were incubated with the Ricinus communis lectin specific for D-galactose residues. The bound lectin was then localized with gold granules labeled with a galactose-terminated glycoprotein (desialylated ceruloplasmin). The galactan was found in the nuclei mainly associated with chromatin and, also, but to a smaller extent, in the cytoplasm and in some vacuoles. The specificity of the method was assessed by marking under the same condition the galactomannan present in the cell wall of the yeast Schizosaccharomyces pombe.


The origin of the nitrogen atom in the thiazole ring of thiamine in Escherichia coli. Methods are described for the isolation and gas chromatographic-mass spectrometric analysis of the 4-methyl-5-beta-hydroxyethyl thiazole moiety of thiamine in microbial cells. Using these methods, it was determined that in Escherichia coli the nitrogen atom in the thiazole ring of thiamine is derived solely from L-tyrosine.


Isolation, purification, and partial chemical characterization of chromium(III) fractions existing in brewer's yeast and Sabouraud's liquid medium. An ethanol extract of brewer's yeast which had been cultivated in a medium containing trivalent 51Cr was analyzed for 51Cr compounds by using petroleum ether extraction, gel filtration, cation and anion exchange chromatography and thin layer chromatography. Similar analytical procedures as for the above analysis were used for studying 51Cr compounds formed in the spent culture medium and in a sterile medium. Several 51Cr fractions were isolated from the three chromium sources, but one anionic 51Cr fraction present in the yeast and in the spent culture medium was not found in the sterile medium. Molecular weight estimations of the 51Cr fractions by gel filtration chromatography showed that the 51Cr ion exchange fractions contained several 51Cr compounds. The molecular weights of these compounds ranged from 150 to 1000 daltons and the molecular weights of 51Cr compounds separated from the yeast were markedly lower than those of the corresponding ion exchange fractions isolated from the culture medium. By using thin layer chromatography it was possible to isolate 51Cr compounds from the main bulk of ninhydrin active impurities. The polarity of all 51Cr compounds was found to be greater than that of most amino acids. The 51Cr compounds isolated from the yeast were mixed with 125I-insulin and incubated, after which the solution was eluted through Sephadex G-50 gel to test if binding had occurred. Elution peaks of 51Cr and 125I-insulin showed that 51Cr compounds were not bound to the insulin.


Insulin action on Escherichia coli. Regulation of the adenylate cyclase and phosphotransferase enzymes. Insulin on Escherichia coli was studied using wild type E. coli B/r and K12 strains and a number of phosphoenolpyruvate phosphotransferase mutants. In vivo, the effects of insulin on the differential rate of tryptophanase synthesis, the rate of alpha-methylglucoside uptake and the rate of growth on glucose were determined in E. coli B/r. In vitro, the effect of insulin on the adenylate cyclase and the phosphotransferase activities was determined using toluenized cell preparations of E. coli B/r, E. coli K12 and phosphotransferase mutant strains. The specificity of insulin action on E. coli was determined using glucagon, vasopressin and somatropin as well as insulin antisera. Results show the specific action of insulin on E. coli, inhibiting tryptophanase induction and adenylate cyclase activity, while stimulating growth on glucose and uptake and phosphorylation of alpha-methylglucoside.


Biphasic saturations of binding proteins can be the result of a competitive inhibition of substrate fixation. A mathematical model is proposed which explains some biphasic saturations of Binding Proteins by their substrates through an effect of a competitive inhibition. The inhibitor can be the substrate itself especially when the retention phenomenon is occuring. This model has been verified with two periplasmic Binding Proteins of Escherichia coli: the Glutamine Binding Protein and the Leucine-Isoleucine-Valine Binding Protein. A significant connection is found between experimental results and the hypothesis.


Finger pulse amplitude and frontalis EMG biofeedback effects of single- and two-system training. The relationship between finger pulse amplitude (PA) and frontalis EMG was studied first by looking at general physiological changes accompanying successful bidirectional PA control. Seven successful subjects were then asked to produce two patterns of PA and EMG (PAincEMGdec and PAdecEMGdec) while receiving both PA and frontalis EMG biofeedback. Results indicate subjects can easily produce the differentiation pattern of PAdecEMGdec but cannot produce the integration pattern of PAincEMGdec. These rather paradoxical results may indicate subjects were using an "attentional" rather than "arousal" strategy for controlling PA and have implications for the use of peripheral vasomotor training as a general relaxation technique.


The relative effectiveness of three techniques to induce the trophotropic response. The purpose of this study was to examine the relative effectiveness of electromyographic biofeedback training (EMG BFT), meditation, and progressive muscle relaxation (PMR) in eliciting a relaxation or trophotropic response as measured by frontalis muscle tension, heart rate, electrodermal response, respiration rate, and skin temperature. Fifty-four college students were randomly assigned to one of five groups: (1) control, (2) placebo control, (3) EMG BFT, (4) meditation, (5) PMR. After baseline measures were obtained subjects were trained in 10 30-minute training sessions and posttested. Comparisons by ANOVAs indicated there was a significant decrease in muscle tension in the EMG BFT and meditation groups and significant decreases in respiration rate in the meditation and PMR groups. No other changes were attributed to treatment.


1/f noise in membranes.

1/f noise in membranes. The present situation of 1/f noise in the passage of ions across membranes is examined. A survey of biological and synthetic membranes is given at which a 1/f frequency dependence has been observed in the spectrum of voltage or current fluctuations. Empirical relations and theories of 1/f noise in membranes are critically discussed.


A contrast variation study of Escherichia coli ribosomes reassembled from protonated and deuterated subunits. Neutron low angle scattering studies on E. coli ribosomes reassembled from protonated and deuterated subunits indicate that the association of the two subunits occurs without major distortion of their shape or modification of the distribution of the protein and RNA components.


Immobilization of immunoglobulins on polystyrene latex beads: characterization by density gradient centrifugation. Isopycnic banding by density gradient centrifugation was used to measure density changes in complexes formed by the immobilization of each of four different immunoglobulins (IgG) (bovine, dog, rabbit, and sheep) on polystyrene latex beads (0.109 +/- 0.0025 micrometer diameter). Subtractive measurements of density changes allowed calculation of the mass of immobilized IgG under varying experimental conditions. The immobilization data were correlated with adsorption isotherms which incorporated charge repulsion forces. The effects of pH and NaCl concentration on the immobilization were studied for the latex-bovine IgG system. It was found that the mass of immobilized immunoglobulins was increased from 10 to 20% by removing the IgG from its isoelectric range.


[Common antigens of stable group A streptococcal L forms and muscle fibers of human myocardium]. The immunofluorescence method was used to demonstrate that the antigens of stable streptococcus L-forms and of the cytoplasmic membrane of human myocardium muscle fibers were common. The common antigen was included into the composition of the surface membrane of the muscle cell adjacent to the sarcolemma, and of the transverse tubule membranes of the sarcoplasma reticulum passing in the Z-band region of the muscle fiber sarcomeres. The reactions is completely prevented by the exhaustion of the anti-serum to the antigen of the L-forms with the homogenate of human myocardium tissues or a suspension of the L-form cultures grown both on meat and on casein media. Exhaustion with tissue homogenate of other organs (the liver) or with concentrated nutrient medium practically failed to influence the extent of this reaction. The authors believe that the commonnes of the antigens of stable L-forms of streptococcus, group A, cultures and of the myocardium could serve as one of the causes of prolonged persistence of L-forms in human and animal organism.


[Age and increase in the sensitivity threshold of the tonic region of the hypothalamic sex center to inhibition by estradiol in female rats]. Estradiol-17 beta was injected into the 3rd ventricle of hemiovariectomized rats to study the sensitivity of the hypothalamic sexual centre. In aged (14--16-month-old) rats the dose of estradiol required for suppression of the compensatory ovarian hypertrophy by 50 and 100 per cent was 4--5 times greater than in young (3-month-old) rats. The results obtained suggest the age-associated increase of the threshold sensitivity of the tonic region of the hypothalamic sexual centre to estrogen suppression. These data can be used for the interpretation of the mechanism of both the age-associated elevation of gonadotropin secretion, and of the reproductive function.


[Insulin-forming activity of monolayer cultures of pancreatic islet cells from cattle fetuses]. The radioimmunological method was applied to the study of insulin content in the growth medium of primary monolayer cultures of bovine fetal pancreatic islet cells grown with usual and increased (300 mg%) glucose content. The latter led to an enhanced insulin secretion. The results of cytological study demonstrated a definite interrelationship between the mitotic activity of culture cells and the intensity of insulin secretion into the medium.


[Assessment of the functional state of the cerebral cortex in rabbits according to electroencephalographic findings using a computer diagnostic system]. Computer estimation of the current functional state of the cerebral cortex in rabbits by the EEG data permitted to distinguish its peculiarities in dark and light adaptation of different duration under the action of nembutal; individual peculiarities of the cerebral cortex activity were revealed in various rabbits. The automatic converter system and that of recording and analysis of multiprocess information (apromin system) and computer diagnostic system (mds) were used in complex for the multistage processing of the multiprocess information. Final results are presented on the plane as points reflecting multidimentional images of the current functional states of the cerebral cortex in animals.


Fetal hemoglobin in paroxysmal nocturnal hemoglobinuria (PNH): evidence for derivation of HbF-containing erythrocytes (F cells) from the PNH clone as well as from normal hemopoietic stem cell lines. The cellular distribution of HbF was studied in nine patients with paroxysmal nocturnal hemoglobinuria (PNH) by measuring the level of HbF and determining the number of HbF-containing red cells (F cells) in whole blood and in the population of normal cells obtained after immune lysis of the abnormal erythrocytes. The amounts of HbF and the F cell frequencies found in the normal red cells were strikingly similar to the values seen in whole blood. The observed frequencies of F cells in normal cells best fitted those expected under the assumption that the F cells arise equally from normal hemopoietic stem cells and from the stem cells with the PNH defect. Since PNH appears to be a clonal hemopoietic stem cell disorder, this evidence argues against a derivation of F cells from distinct pluripotent stem cell lines.


Primary systemic amyloidosis: comparison of melphalan and prednisone versus placebo. Satisfactory treatment for primary amyloidosis does not exist. Because the amyloid fibrils consist of a portion of a monoclonal light chain, it appears reasonable to treat amyloidosis with alkylating agents that are effective against the plasma cells that synthesize monoclonal light chains. Fifty-five patients with primary systemic amyloidosis were randomized (double blind) to melphalan-prednisone or placebo. In comparison with the placebo group, patients given melphalan-prednisone were able to continue on treatment for a longer time and to receive larger doses before the code was broken. Among this group, the nephrotic syndrome disappeared in two patients and urinary excretion of protein was reduced by more than 50% in eight others. Of 13 patients who received melphalan-prednisone for more than 12 mo, 6 improved, 3 were stable, and 4 had progression of disease. Survival did not differ significantly between the groups.


Global capacity reduction and schizophrenia. This paper reviews the evidence for a specific deficit in the flow of information through the schizophrenic processing system. It is suggested that there is little to support the notion of a single defective mechanism underlying schizophrenic cognitive dysfunction. The lack of a valid and reliable metric for measuring degree of difficulty across tasks and a lack of concern with matching tasks for attentional processing load, detracts from the usefulness of available results. The applicability of a global limited capacity model of selective attention to schizophrenic information processing is discussed as the most appropriate means of systematizing the wealth of published empirical data.


Vitamins and sport.

Vitamins and sport. In a double blind test on 40 men and women high performance fencers the influence of a multivitamin-electrolyte-preparation on reaction time, hit-frequency and neuromuscular irritability was determined. Evidence of B1, B2 and B6-deficiencies up to 70% were found. After application of the preparation an increase of performance of 3% could be observed. The tests were carried out by means of coenzymatic activation of the respective vitamins in the haemolysate of erythrocytes. The results are explained by the role of the vitamins in the human organism and by the special conditions under which endurance athletes have to perform: high turn-over rate of the energy metabolism, an increase of the body core temperature and thus increased sweat loss, factors leading to deficiencies of the water soluble B-vitamins.


Topographic projections of substance P and GABA pathways in the striato- and pallido-nigral system: a biochemical and immunohistochemical study. The topographical projections of substance P pathways from the caudateputamen and globus pallidus to the pars compacta and pars reticulata of the substantia nigra have been investigated in the rat using immunohistochemical and radioimmunoassay techniques and compared with the projections of GABA nergic striatal neurones. Unilateral vertical knife cuts through the anterior and posterior striatum have shown the majority of substance P-containing neurones which project to the substantia nigra to originate in the most rostral part of the caudate-putamen. This projection appears to innervate the pars reticulata and pars compacta of the substantia nigra to a similar extent. A separate projection of substance P-containing neurones to the substantia nigra appears to originate in the globus pallidus. Undercutting the cerebral cortex which overlies the corpus striatum did not affect the substance P content of the globus pallidus or substantia nigra. However, there appears to be an additional substance P projection from the basal ganglia to the entopeduncular nucleus. In contrast, GABA-containing neurones which project to the substantia nigra are mainly located in more caudal parts of the caudate-putamen and in the globus pallidus. There is a marked differentiation in the region of the substantia nigra innervated by GABA cells originating in the rostral and caudal parts of the corpus striatum. Rostrally situated neurones project almost exclusively to the pars reticulata, while neurones in the caudal part of the caudate-putamen and globus pallidus project to both the pars compacta and pars reticulata. These results suggest that there is a partial topographical separation of the sites of origin of substance P- and GABA-containing neurones which project to the substantia nigra.


The central and peripheral ends of the substance P-containing sensory neurones in the rat trigeminal system. The distribution of substance P (SP) immunoreactivity in the spinal nucleus of the rat trigeminal nerve and in the skin of the lower lip was examined following (a) unilateral electrolytic lesions of the trigeminal ganglion, (b) trigeminal rhizotomy, and (c) unilateral interruption of the mental nerve, the sensory branch of the trigeminal nerve innervating the lower lip. A marked depletion of SP immunoreactivity in the ipsilateral trigeminal spinal nucleus followed lesions of the trigeminal ganglion or rhizotomy. The reticular formation ventral and medial to the spinal nucleus showed a small decrease in SP immunofluorescence on the operated side. Some loss of SP immunoreactivity was observed in the skin of the lower lip following ganglionectomy or rhizotomy. After sectioning the mental branch SP-immunofluorescent fibres of the skin of the lower lip disappear completely on the denervated side. It was concluded that some trigeminal ganglion neurones store, and might release, SP at their axon terminals in the medulla oblongata and at their sensory terminals in the skin.


Changes in intracellular organization of tubulin and actin in N-18 neuroblastoma cells during the process of axon extension induced by serum deprivation. Specific antibodies against actin and tubulin have been used to follow the distribution and organization of actin and tubulin containing structures in N-18 neuroblastoma cells induced to sprout axons. Immunofluorescence microscopy shows that during the time of axonal sprouting microtubules converge into growing processes forming dense bundles in which individual microtubules cannot be resolved. In the growth cone where individual fluorescent fibers can again be distinguished microtubules seem to be excluded from the very margin. Actin is predominantly located at the cell periphery both in cell bodies and in cell processes. It appears to be present in areas of high surface motility and is especially abundant at the tip of the growth cone.


VIP-, enkephalin-, substance P- and somatostatin-like immunoreactivity in neurons intrinsic to the intestine: immunohistochemical evidence from organotypic tissue cultures. Small intestine from 18-day fetal mice grown for 3 weeks in organotypic tissue culture was found to contain numerous VIP, enkephalin, substance P and some somatostatin immunoreactive nerve fibers. Since these cultures should be devoid of all afferent or other extrinsic neuronal inputs, it is concluded that there are VIP, enkephalin, substance P and somatostatin containing neurons intrinsic to the intestinal wall. However, all 4 peptides may also be present in neurons originating outside the gastrointestinal tract as well as in the intrinsic neurons.


Levels of coenzyme A--glutathione mixed disulfide in Escherichia coli. The pool of coenzyme A--glutathione mixed disulfide (CoASSG) rapidly increased 2.0 times in response to oxygen starvation and 1.5 times in response to glucose starvation but did not change following ammonia starvation. The increase in the CoASSG pool resulted from an increase in the CoASSG fraction of the CoA pool from 42 to 66--93%. Fluoride, cyanide, chloramphenicol, and rifampicin all caused similar increases. Aerobic growth on fermentable sugars resulted in CoASSG making up 40--55% of the CoA pool while growth on nonfermentable carbon sources or anaerobic fermentation resulted in CoASSG replacing acetyl CoA and free CoA to make up 85--95% of the CoA pool. The CoASSG:ATP ratio varied inversely with the growth rate in two groupings of carbon sources made up of either fermentable or nonfermentable molecules. Cultures grown aerobically on fermentable sugars exhibited a lower CoASSG:ATP ratio reflecting the lower proportion of CoASSG in the CoA pool.


Metabolism of delta24-sterols by yeast mutants blocked in removal of the C-14 methyl group. We have investigated the metabolism of exogenously provided delta24-sterols by whole cell cultures of a polyene-resistant mutant (D10) of Candida albicans blocked at removal of the C-14 methyl group. Comparison of the relative efficiencies of transmethylation at C-24 of selected sterol substrates revealed the following substrate preferences of the Candida delta24-sterol methyltransferase (EC 2.1.1.41): zymosterol greater than 4alpha-methylzymosterol greater than 14alpha-methylzymosterol. Exogenous 4,4-dimethylzymosterol was not transmethylated by mutant D10. Incorporation of the 14C-labelled methyl group of S-adenosyl-L-[methyl-14C]methionine into the sterols of a D10 culture preloaded with zymosterol indicated that zymosterol was a better (40 X) substrate than endogenous lanosterolmfeeding zymosterol to D10 and a polyene-resistant strain of Saccharomyces cerevisiae (Nys-P100) that was also blocked at removal of the C-14 methyl group gave 24-methyl sterols possessing delta22 and ring B unsaturation. Mutant D10 was able to produce ergosterol from zymosterol whereas Nys-P100 produced ergosta-7,22-dienol. When grown in the presence of 3 micrometer 25-aza-24,25-dihydrozymosterol, a known inhibitor of the delta24-sterol methyltransferase, Nys-P100 accumulated 14alpha-methylzymosterol, a minor metabolite in this mutant under normal growth conditions and hitherto unidentified as a yeast sterol.


Phosphorylation of adenosine 5'-monophosphate by a dialyzed cell-free extract from Escherichia coli. A cell-free extract of Escherichia coli, even after exhaustive dialysis, was found capable of phosphorylating adenosine 5'-monophosphate (AMP) to adenosine 5'-diphosphate (ADP) and adenosine 5'-triphosphate (ATP). Centrifugation at 100 000 g for 3h sedimented most of the capacity to phosphorylate AMP to ATP, while the supernatant retained a significant capacity to phosphorylate AMP to ADP. The pellet contained a greater amount of phosphate polymers (which were neither DNA, RNA, nor proteins) than did the supernatant. The addition of authentic inorganic polyphosphates to the supernatant restored the phosphorylating capacity of the original extracts. It is concluded that the observed phosphorylation is partly due to inorganic polyphosphate.


The effect of adsorbant and anti-inflammatory drugs on secretion in ligated segments of pig intestine infected with Escherichia coli. Four adsorbant drug preparations, Kaopectate, colloidal Attapulgite, noncolloidal Attapulgite and Pepto-bismol were investigated for their effects on fluid accumulation in ligated segments of pig intestine inoculated with enteropathogenic Escherichia coli. Two anti-inflammatory drugs. aspirin and methylprednisolone, and two antibiotics, lincomycin and polymyxin B, were also tested. All the drugs except the two anti-inflammatory products were given by injection into the lumen of the intestine. Aspirin was given orally and methylprednisolone was given intramuscularly. The antibiotics were tested at levels at which they had no significant antibacterial effect in in vitro tests. The adsorbant drugs colloidal Attapulgite and Pepto-bismol were shown to be effective in reducing fluid accumulation in ligated segments of pig intestine infected with enteropathogenic E. coli. In the case of Peptobismol this effect was associated with an antibacterial effect as well as an antitoxic effect, probably due to its adsorbant properties. It is possible that an aspirin-like effect in the gut due to the active ingredient bismuth subsalicylate may have contributed to the effectiveness of Pepto-bismol. Colloidal Attapulgite was demonstrated to have an antitoxic effect but did not have an antibacterial effect. In high doses, the anti-inflammatory drugs acetylsalicylic acid and methylprednisolone were marginally effective in reduction of fluid accumulation in the same test system. Lincomycin was shown to reduce intestinal fluid secretion, whereas polymyxin B had no effect.


Demonstration of parvovirus in Canadian swine and antigenic relationships with isolates from other countries. A Canadian isolate of porcine parvovirus, isolated from cultured pig thyroid cells, was shown to be antigenically indistinguishable from a British (59e/63) and a German (G10/1) strain when treated by the modified direct complement-fixation, the hemagglutination-inhibition and the fluorescent antibody tests. These tests also revealed that antibodies to parvoviruses were detectable in a large proportion of the conventionally raised pigs in the provinces of Quebec and Ontario. Cell cultures, prepared from tissues collected in a slaughterhouse, were often found to be infected with parvovirus. In cell cultures the infection was demonstrated more effectively by immunofluorescence than by the hemagglutination test.


Germination and outgrowth of Schizosaccharomyces pombe ascospores isolated by Urografin density gradient centrifugation. A simple method for the isolation of single ascospores of the fission yeast Schizosaccharomyces pombe was examined. Single spores in the 7-day-old sporulating culture of a homothallic strain were separated from remaining vegetative cells by isopycnic centrifugation in the linear gradient from 10 to 60% of Urografin solution at 700 X g for 20 min. Protein content of isolated spores was very low as compared with that of vegetative cells. The isolated spores germinated through the following steps when cultured in a liquid medium at 25--35 degrees C; loss of refractility (darkening) under a phase-contrast microscope, spherical growth (swelling), emergence of germ tubes, elongation of germ tubes, cell plate formation, and cell separation. The absorbance at 650 nm of the spore suspension initially decreased, accompanied by darkening of spores, and then increased with spherical growth. The germination rate of isolated spores reached almost 100%.


Effect of heat stable and heat labile Escherichia coli enterotoxins and cholera toxin in combination with theophylline on unidirectional sodium and chloride flux in the small intestine of weanling swine. The effect of heat stable and heat labile Escherichia coli enterotoxins or cholera toxin in combination with theophylline on net water, sodium and chloride and unidirectional sodium and chloride fluxes was examined in acute isolated loops of jejunum of weanling swine. The effect of heat stable enterotoxin in combination with theophylline was determined in loops located in the proximal jejunum, while combinations of theophylline and either heat labile enterotoxin or cholera toxin were studied in the distal jejunum. In each situation the addition of theophylline resulted in an additive rather than a synergistic increment of intestinal secretory activity. This study implies that the intestinal adenyl cyclase system and enterotoxin induced intestinal secretion may not be directly related in the swine small intestine.


Postbacterial sepsis and disseminated candidiasis. The effect of gram-negative sepsis (Escherichia coli) on the capacity of polymorphonuclear leucocytes (PMN) to phagocytize and kill Candida albicans was assessed. The PMN's from septic dogs phagocytized C. albicans as well as PMN's from non-septic dogs. The PMN's from septic dogs that phagocytized C. albicans underwent a spontaneous lysis at a much higher rate than PMN's from non-septic dogs. A functional difference in PMN's from normal and septic dogs is indicated.


A taxonomic study of the Spirillum lipoferum group, with descriptions of a new genus, Azospirillum gen. nov. and two species, Azospirillum lipoferum (Beijerinck) comb. nov. and Azospirillum brasilense sp. nov. Sixty-one strains of the root-associated nitrogen fixer Spirillum lipoferum exhibited a similar morphology in peptone--succinate salts medium: vibrioid cells having a diameter of 1.0 micrometer. When grown in broth the cells had a single polar flagellum, but when grown on agar at 30 degrees C lateral flagella of shorter wavelength were also formed. The DNA base composition was 69--71 mol% guanine + cytosine when determined by thermal denaturation. DNA homology experiments indicated the occurrence of two distinct but related homology groups: 46 strains were in group I and 15 strains were in group II. Group II strains were distinguished by their ability to use glucose as a sole carbon source for growth in nitrogen-free medium, by their production of an acidic reaction in a peptone-based glucose medium, by their requirement for biotin, and by their formation of wider, longer, S-shaped or helical cells in semisolid nitrogen-free malate medium. The results indicate that two species exist, and on the basis of their characteristics it is proposed that they be assigned to a new genus, Azospirillum. Strians belonging to group II are named A. lipoferum (Beijerinck) comb. nov., while those belonging to group I are named A. brasilense sp. nov. Strain Sp 59b (ATCC29707) is proposed as the neotype strain for A. lipoferum, and strain Sp 7 (ATCC 29145) is proposed as the type strain for A. brasilense.


Flunisolide nasal spray in the treatment of perennial rhinitis. In an 8-week double-blind study of the clinical effectiveness of flunisolide, a new synthetic corticosteroid intended for topical use, 73 persons with perennial rhinitis received 300 microgram/d of either flunisolide or placebo. The flunisolide group showed a significantly greater reduction than the placebo group in the daily duration of sneezing, stuffy nose, nose blowing and postnasal drip, but not of runny nose, and a significantly greater reuction in the severity of all these manifestations. Substantial or total control of manifestations was reported by 73% of the flunisolide group and 25% of the placebo group, a significant difference. Careful monitoring of the plasma  cortisol concentration revealed no evidence of adrenal suppression, and in no patient did the value fall below normal. The lack of adrenal suppression may be due to the very small amount of corticosteroid administered and the rapid metabolism of flunisolide into a relatively inactive metabolite.


Non-hodgkin's lymphomas in Saskatchewan: a clinicopathologic study. In a retrospective clinical study of 208 previously untreated persons with non-Hodgkin's lymphomas the disorders were classified and staged according to the histopathologic criteria of Rappaport, Winter and Hicks and the Ann Arbor clinical staging classification.Nodular types constituted 22% and diffuse types 78% of the lymphomas. The nodular lymphomas were slightly more common in females and were clustered in the age range 30 to 90 years. The diffuse lymphomas were slightly more common in males; the age distribution was bimodal, with one peak in the age range 10 to 19 years and the other in the age range 60 to 69 years, but when the age distribution of the general population in which the lymphomas occurred was taken into account, the incidence of these lymphomas was found to be significantly higher (P < 0.001) in persons more than 69 years of age than in those 40 to 69 years of age.SURVIVAL CORRELATED WITH HISTOPATHOLOGIC TYPE: persons with nodular (follicular) lymphomas and diffuse lymphocytic well differentiated lymphomas had a significantly greater survival (P < 0.05) than those with other diffuse lymphomas. No significant difference in survival was noticed between persons with nodal and extranodal lymphomas.While Rappaport and colleagues' criteria are still very useful, it is important to recognize the nodular lymphoma as a specific entity requiring generally different management from diffuse lymphomas. Appreciation of the different biologic behaviour of the various lymphomas is important to clinicians planning therapy.


Complications of treatment encountered in lymphoma-leukemia long-term survivors. Potential posttreatment complications for patients with the lymphomas and leukemia include: 1) impairment of growth and development in children, 2) CNS disturbances encompassing psychologic, intellectual and neurologic expressions, 3) gonadal effects--endocrine, reproductive, teratogenic and genetic, 4) dysfunction of other organs and structures, such as the thyroid, lung and heart and 5) oncogenesis. Many of these adversities are occasioned by radiation therapy, but chemotherapy also can be responsible for some long-term deleterious consequences. The results of combined chemo- and radiaton therapy are becoming better understood, and require further elucidation because earlier stages of the disease are being managed by combined chemo- and radiation therapy regimens.


The immunological classification of acute lymphoblastic leukemias. The main methods used in order to assess the immunological markers on the blast cells from patients with acute lymphoblastic leukemia (ALL) are critically evaluated. The classification of ALL according to the phenotype of the leukemic cells and their presumed origin is reviewed. The correlations with clinical features and prognosis, as well as their possible therapeutic implications, are discussed.


Acute lymphoblastic leukemia: treatment. Over the past thirty years, major progress has been achieved in the treatment of ALL. Many of the concepts, definitions, and principles of chemotherapy have and continue to be derived from studies in ALL. Major and continuing progress is ongoing for the various categories of treatment; that is, remission induction, treatment at sites of high risk for relapse (e. g., pharmacologic sanctuaries), cytoreductive therapy during complete remission, and duration of treatment. Rapid improvement in our understanding of the pathogenesis of ALL and particularly the identification of immunologic and prognostic subcategories of ALL have major therapeutic implications which are in process of being realized. Current research is focused on the development of new chemotherapeutic agents, the more rational basis for the employment of chemotherapeutic agents in combination (cytokinetic, pharmacologic, and related studies), a better definition of the host-tumor relationship particularly with respect to immunologic response and iatrogenic manipulation of such responses, and in the area of supportive care and bone marrow transplantation.


The preleukemic syndrome (hemopoietic dysplasia). Retrospective analysis of patients with acute myelomonoblastic (or nonlymphocytic) leukemia has helped to clarify the early stages of the evolution of this disease, and a clinical picture has emerged that is sufficiently discriminating to permit prospective observations. The clinical and laboratory features of the hematologic disorder preceding overt, blast-cell leukemia (i.e., the "preleukemic syndrome" or "hemopoietic dysplasia") are described, and diagnostic criteria and approaches to management are considered. This hematologic syndrome appears to be a stage in a multiphasic panmyelopathy that involves all products of the marrow stem cell. Available data indicate that most of these patients develop acute leukemia within 24 months of the onset of the hematologic abnormalities, but longer courses are not uncommon. Prospective documentation in a large group of patients is needed before the natural course can be accurately defined; such studies are now being planned.


Immunologic aspects and pathology of the malignant lymphomas. Malignant lymphomas have traditionally been classified on solely morphologic grounds. With new immunologic and cytochemical techniques, it has been possible to characterize normal cells of the T-lymphocytic, B-lymphocytic, and monocyte-macrophage systems. Application of these methodologies to malignant lymphomas has established their nature as neoplasmas of the immune system. Within the B-lymphocytic system it is possible to identify subpopulations responsible for Burkitt's tumor, follicular (nodular) lymphomas, lymphocytic lymphomas of intermediate differentiation and well differentiated lymphocytic lymphomas. The T-lymphocytic system includes lymphoblastic lymphomas, mycosis fungoides, and Sezary's syndrome. Large cell lymphomas are diverse but the majority are tumors of transformed lymphocytes, usually of the B-lymphocytic system. The precise nature of the neoplastic cells of Hodgkin's disease, i.e., Reed-Sternberg cells and their mononuclear counterparts, has not yet been established. Despite previous suggestions of a B-lymphocytic or T-lymphocytic origin, recent studies utilizing in vivo cultivation have strongly suggested derivation from the monocyte-macrophage system.


Hairy cell leukemia: an unusual lymphoproliferative disease: a study of 24 patients. A laboratory and clinical evaluation of 24 patients with hairy cell leukemia was carried out over a 23-month period. Most patients had splenomegaly without adenopathy or pancyotpenia. Nine of the patients had undergone splenectomy prior to referral; their median WBC count was 6600/mm3. The median WBC count for the 14 patients who had no prior therapy was 3550/mm3, and their median platelet count was 80,500/mm3. Spleen weights ranged from 618 to 3780 g; there appeared to be no relationship between the size of the spleen and the response in the blood counts after splenectomy. Four patients in whom the majority of the WBC were hairy cells underwent splenectomy, which produced no real change in their WBC count; however, there was improvement in the platelet count in three. In contrast, the presence of leukopenia with a low percentage of hairy cells predicted a beneficial response to splenectomy. The study of surface immunoglobulins (SIg) in 16 patients demonstrated that resynthesis had occurred in each case. Phagocytosis of zymosan was studied in 15 patients; in 8 of these, 25% or more of the hair cells were capable of phagocytosis; in 6 others, 0--9%; and in one, 13%. The resynthesis of SIg is a feature usually associated with B-lymphocytes, but the phagocytosis of zymosan is not. Thus, the existence of either a spectrum of functional capabilities of hairy cells or several distinct subtypes is suggested by these data. Platelet aggregation with epinephrine was abnormal in 7 of 14 patients studied but there were no clinically significant bleeding problems. A chromosome abnormality was present in 2 of the 19 patients from whom adequate samples were obtained; the abnormality probably involved chromsome 12 in both patients as well as absent Y and was associated with a rapidly progressive clinical course. The presence of a predominant number of hairy cells with a normal or increased peripheral blood WBC count or of a chromosomal abnormality suggests that splenectomy might not be beneficial as the initial therapy and that chemotherapy should be considered.


The chemotherapy of Hodgkin's disease: past experiences and future directions. From the standpoint of chemotherapy, the first progress in the treatment of Hodgkin's disease was the identification of the activity of nitrogen mustard in the 1940's. The initial antitumor effect of the drug created a great excitement. However, when all patients later relapsed, there was subsequent dejection and skepticism about the utility of drug therapy. Fortunately, in the 1950's and 1960's, the development of other effective agents (vinca alkaloids, corticosteroids, and methylhydrazines) in conjunction with the elucidation of the principles of combination chemotherapy led to a marked increase in the antitumor response rate of patients with Hodgkin's disease. The value of many of these drug combinations remains under study. Nonetheless, approximately 75% of all patients with advanced Hodgkin's disease treated today with combination chemotherapy can achieve a complete remission. In addition, over half of these remain disease-free long enough to be considered cured. The development of effective treatment,  both local (radiotherapy) and systemic (MOPP chemotherapy), has given the clinical investigator the tools to complete, in the 1970's, the therapeutic experiments necessary to refine both the interrelationship between the treatments and their impact upon the natural history of Hodgkin's disease.


The need for a mammalian test system for mutagens: action of some reducing agents. Reducing agents and cysteine, cysteamine, glutathione, ascorbic acid and H2O2 with and without the addition of Cu2+ did not increase significantly the frequency of mutations in the Salmonella test at non-toxic concentrations but triggered a marked DNA repair synthesis and induced a relatively high frequency of chromosome aberrations in cultured mammalian cells. Both latter effects were reduced by the addition of catalase to solutions of the reducing agents plus Cu2+. To avoid 'False Negatives' in mutagenicity screening the use of several test subjects including mammalian cells seems to be required.


Controlled trial of methotrexate and Bacillus Calmette-Guérin therapy for advanced head and neck cancer. Thirty-eight patients with advanced, inoperable squamous cell carcinoma of the head and neck were randomized to receive methotrexate alone or methotrexate with Bacillus Calmette-Guérin. The response rates with methotrexate (3 of 19) and methotrexate plus B. Calmette-Guérin (4 of 16) were similar, as was the duration of response and survival of the two groups. The results of in vitro immunological studies of lymphocytes were assessed. Marked weight loss, poor performance status, and distant metastases were the most important prognostic factors. The presence of anergy was significantly correlated with weight loss. This study also indicated that a large tumor burden is a frequent occurrence in advanced head and neck cancer and may account for the lack of efficacy of B. Calmette-Guérin.


Clinical trial of peptichemio in solid tumors of childhood. Peptichemio (PTC) was utilized in various advanced and disseminated tumors in children. With the schedule that was used, toxicity was mild. Immediate side effects included anaphylactoid reactions which required close supervision of the patient during infusion of the drug. PTC proved efficacious in advanced neuroblastoma. In patients with bone metastases, PTC has a prompt analgesic effect which seems to occur independently of any antitumor action.


Clinical trial of rubidazone in solid tumors and malignant lymphomas. Rubidazone, a semisynthetic daunorubicin derivative, was administered to 33 patients with advanced solid tumors or malignant lymphomas. The doses used were 200, 150, or 100 mg/m2 repeated at 3-week intervals. The antitumor effect was very modest. Partial remission was obtained in only one patient and stable disease was observed for periods of 4--18 weeks in seven additional patients. The major toxic effects were gastrointestinal and myelosuppressive. In two patients, cardiac toxic effects were noted and one patient had an anaphylactic-like reaction. No clinical cross-resistance seemed to exist between rubidazone and adriamycin.


Combination chemoimmunotherapy for extensive non-oat cell lung cancer. In a prospective randomized trial, sc Corynebacterium parvum (C. parvum) immunotherapy did not significantly affect the responses and survival of 49 non-oat cell lung cancer patients receiving isophosphamide and adriamycin chemotherapy. Remissions (tumor regression greater than 50%) were seen in five of 23 patients receiving an intensive C. parvum schedule and in three of 26 patients receiving a nonintensive C. parvum schedule (22% versus 12%). Median survival was 20 weeks for patients given intensive C. parvum and 23 weeks for patients given nonintensive C. parvum. This study did demonstrate the importance of pretherapy immunocompetence, performance status, and weight loss as predictors for survival. Weight loss was the most significant prognostic factor. Performance status was closely associated with weight loss but skin reactivity to dermatophytin predicted independently and was the second most important prognostic characteristic.


Chemoimmunotherapy for disseminated malignant melanoma: a prospective randomized study. Twenty-eight patients with disseminated malignant melanoma were treated with DTIC (250 mg/m2 iv, Days 1--5) and actinomycin D (0.5 mg/day iv, Days 1--5) at 5-week intervals. Patients were randomly allocated to receive no immunotherapy or immunotherapy consisting of methanol extracted residue of bacillus Calmette-Guérin (0.5 mg intradermally; 100 microgram in five separate sites) every 5 weeks, concomitant with chemotherapy. Of these 28 evaluable patients, 13 received chemoimmunotherapy with one complete response (CR) and 15 received chemotherapy alone with one CR. Both responses occurred in lymph node metastases. No partial responses were seen.


Phase II study of vinblastine, methyl-CCNU, and medroxyprogesterone in advanced renal cell cancer. One hundred and sixty-five patients with advanced renal cell cancer were evaluable for combination or single-agent therapy with methyl-CCNU, vinblastine, and medroxyprogesterone. A low order or response was observed, and these agents were not proven effective as treatment for metastatic renal cell cancer. Performance status and a relatively long symptom-free interval from primary tumor to metastatic disease were found to be the most prognostically significant factors for survival.


Piperazinedione in patients with advanced malignant melanoma: a Southwest Oncology Group study. Piperazinedione was administered in doses of 9 or 12 mg/m2 by iv infusion every 3 weeks in 28 patients with previously treated malignant melanoma. Of the 25 evaluable patients, 72% had drug-induced toxicity: 40% had leukopenia, 56% had thrombocytopenia, 40% had anemia, and 16% had nausea and vomiting. None of these patients had partial remission, two had stable disease, and the remaining 23 had definite progression of their disease in spite of adequate trial with this agent.


Results of treating stage I and II carcinoma of the breast with primary radiation therapy. Eighty patients with stage I and II carcinoma of the breast were treated with primary radiation therapy between July 1968 and December 1975 at the Joint Center for Radiation Therapy. The median followup time was 41 months. Only one local recurrence was observed. The survival at 5 years was 88% for stage I and 72% for stage II. The relapse-free survival at 5 years was 91% for stage I and 56% for stage II. Of breasts evaluated for cosmetic results, 66% were judged good or excellent by physicians and 81% were judged good or excellent by patients. One patient developed a pericardial reaction related to treatment which required hospitalization. Other complications were mild and cleared without additional therapy.


Cell kinetics and chemotherapy: a critical review. The paper reviews methods of studying cell kinetics in man, cell population kinetics of human tumors and bone marrow, drug interactions and the cell cycle, and possible applications to chemotherapy. The conclusions drawn are: (1) Cell cycle time and S-phase duration for proliferating granulocyte precursors in human bone marrow are poorly defined but are probably shorter than median values for most human tumors, including leukemia. (2) Most drugs have greater toxicity for cycling cells and some variation in toxicity at different phases of the cell cycle. There is a special need for chemotherapy directed at slowly proliferating and hypoxic tumor cells. (3) Pretreatment indices of tumor cell kinetics are of little value in choosing drugs or in predicting response. (4) Experiments in animals have demonstrated that therapeutic index may depend on schedule. Knowledge of cell kinetics in animals rarely allows prediction of the optimal schedule and is unlikely to do so in man. Optimal schedules in mice are not directly relevant to man. (5) Measurement of tumor labeling index or DNA histogram by flow microfluorimetry to detect cell synchrony is of little benefit in scheduling if concurrent changes in bone marrow are ignored; these methods are invalid at short intervals after treatment because surviving clonogenic cells are indistinguishable from a larger number of drug-damaged cells prior to their lysis. (6) The major factor determining the outcome of chemotherapy is the availability of drugs with activity for the tumor and acceptable host toxicity. Claims that complex schedules using several drugs are effective  because of synchrony or kinetic differences of tumor and normal tissue are at present unsubstantiated.


Phase I study of hycanthone.

Phase I study of hycanthone. Hycanthone was given to 15 patients with metastatic cancer in order to determine the maximum tolerable dose. The drug was administered in 5-day courses at 3-week intervals. The starting dose was 30 mg/m2/day and the highest dose level reached was 90 mg/m2/day. The most common (13 patients) side effect was nausea and/or vomiting. The dose-limiting toxicity was toxic hepatitis manifested as elevation in serum transaminases in eight of 15 patients and an increase in serum bilirubin in three patients. Hepatotoxicity was dose-related and was observed in two of 25 courses given at the dose level of less than or equal to 70 mg/m2 compared to seven of nine courses given at the dose level of greater than or equal to 80 mg/m2. Because of an unacceptable incidence of hepatotoxicity at higher doses, 70 mg/m2/day x 5 appears to be a safe dose for phase II studies.


Phase II trial of a combination of cyclosphamide, vincristine, and methotrexate in advanced colorectal carcinoma. Twenty patients with advanced colon cancer were treated with a combination of low-dose weekly cyclophosphamide, vincristine, and methotrexate (COM) after demonstrated progression of disease with 5-fluorouracil (5-FU). Four patients (20%) had partial responses with a median survival of 15 months, six (30%) had disease stabilization for 3--16 months (median survival, 8 months), and ten progressed with a median survival of 5.5 months. All four partial responses and five of six stabilizations occurred in patients with a performance status of 0 or 1. COM appeared well-tolerated and at least as effective as alternative secondary chemotherapy following 5-FU failure.


Phase I--II study of intermittent bolus administration of DTIC and actinomycin D in metastatic malignant melanoma. Twenty-six patients with disseminated malignant melanoma were treated with intermittent bolus DTIC and actinomycin D in an escalating dose schedule, starting at 650 and 1 mg/m2 respectively. Courses were repeated at 3--4-week intervals. Twenty four patients were  evaluable for toxicity and 22 were evaluable for response. Two patients (9%) had a complete remission lasting 7+ and 14 months, and three patients (14%) had a partial remission lasting 2+, 5+, and 14+ months. Nausea and vomiting, lasting 24 hours, was observed in 88% of patients, while diarrhea was noted in 17%. Stomatitis and alopecia were less frequently observed. All responses occurred at nonmyelosuppressive doses and in patients with visceral-predominant metastases. This schedule offers the patient the convenience of single-day treatment and less prolonged gastrointestinal intolerance. Further evaluation of this drug combination and schedule would appear to be indicated.


Effect of primary treatment modality on the metastatic pattern of mammary carcinoma. In animal tumor systems, all three major treatment modalities, surgery, radiotherapy, and chemotherapy, may increase the incidence of metastases in the presence of circulating viable tumor cells. In breast cancer patients, selected studies can be found which report an increased incidence of metastases after surgery, radiotherapy, or chemotherapy, but these effects appear to exert little influence on overall survival. Caution is advised in using systemic therapy prior to effective primary tumor cytoreductive treatment. Clinical trials in advanced local disease should be done to test this concern. Minimal surgery, loco-regional radiotherapy, and effective adjuvant systemic therapy may result in the improved survival of patients with breast cancer with minimal functional or cosmetic impairment.


5-fluorouracil, methyl-CCNU, adriamycin, and mitomycin C in the treatment of advanced gastric cancer. Eighteen previously untreated patients with advanced nonresectable adenocarcinoma of the stomach were treated with a four-drug regimen consisting of 5-fluorouracil, methyl-CCNU, adriamycin, and mitomycin C. Objective responses occurred in only two patients (11%); the majority of patients had stable disease for greater than or equal to 8 weeks (61%). The median survival of 27 weeks was similar to that reported in other combination chemotherapy trials. Numerous staging and restaging procedures and strict response criteria were utilized and each was evaluated for its effectiveness in following responses. None of these parameters was useful for evaluating response of intra-abdominal tumor. We conclude that this four-drug combination is no better than previously described two-drug regimens.


Phase II clinical trial with high-dose methotrexate therapy and citrovorum factor rescue. One hundred and thirty-four patients with advanced malignant disease were treated with 496 infusions of high-dose methotrexate (HD-MTX) followed by citrovorum factor rescue. Most patients had failed to respond to previous combination chemotherapy. The overall response rate was 29% with 33 partial responses and six complete responses observed in patients with a variety of tumors. Plasma MTX levels were monitored in all patients during each course of therapy in order to identify those patients with delayed plasma MTX clearance. Patients with abnormally slow rates of plasma MTX decay received escalated doses of citrovorum factor rescue in order to prevent drug-induced toxicity. In general, during this study HD-MTX was well-tolerated. Because serious toxicity was neither frequent, severe, nor unpredictable, its use was not limited. HD-MTX should now be evaluated in well-designed controlled clinical trials to compare its antitumor activity to that of conventional- or standard-dose MTX regimens in diseases where HD therapy appears to have efficacy.


Phase I trial of vindesine in patients with advanced cancer. A phase I evaluation of vindesine was carried out in 69 adult patient with advanced malignancies. Two escalating dose schedules were explored: (a) a single dose every 7--14 days, and (b) daily injections X 5--10 days as tolerated. The main toxic effects were myelosuppression, alopecia, paresthesia, asthenia, myalgia, and hyporeflexia. Antitumor activity was seen during this phase I study in patients with leukemia, lymphoma, and testicular neoplasms. Disease oriented phase II trials of 3--4 mg/m2 every 7--14 days or 1.3--2.0 mg/m2/day X 5--7 days every 3 weeks would be appropriate.


Treatment of adult acute lymphoblastic leukemia with cytosine arabinoside, vincristine, and prednisone. Thirteen adults with acute lymphoblastic leukemia were entered into a protocol in which cytosine arabinoside infusion was added to vincristine and prednisone as remission induction and periodic intensification therapy. Central nervous system prophylaxis consisting of cranial irradiation and intrathecal methotrexate was given, and all patients received continuous oral 6-mercaptopurine and methotrexate as maintenance treatment. Myelotoxicity was severe during induction, with prolonged granulocyte and platelet count nadirs noted. Nine of 13 patients (69%) obtained a complete remission and one (8%) obtained a partial remission. The median duration of complete remission was 38.1 weeks. It was concluded that cytosine arabinoside in combination with vincristine and prednisone is an effective but toxic antileukemic regimen which did not produce a major improvement in remission duration.


Dissecting the hematopoietic microenvironment. III. Evidence for a positive short range stimulus for cellular proliferation. Experiments were carried out with the intent of defining the nature of the microenvironmental defect which severely limits erythropoiesis in the spleen of the S1/S1d mouse. Chimeric spleens, half S1/S1d and half congenic +/+, supported erythropoiesis in the +/+ region but not in the S1/S1d region, regardless of which genotype was the irradiated, marrow-injected host animal. Implants of normal marrow stroma within the spleens of irradiated S1/S1d mice also supported normal proportions of erythrocytic and granulocytic hematopoiesis. Implants of normal spleen stroma, particularly the capsular portion, into unirradiated s1/s1d mice stimulated erythropoiesis originating from S1/S1d stem cells within and in the immediate vicinity of the implant. The evidence suggests a short range, stromal erythropoietic stimulatory factor which is lacking in the S1/S1d.


Analysis of the progression of meiosis in dispersed rat testicular cells by flow cytofluorometry. Initiation and progression of meiosis was followed in dispersed rat testicular cells by flow cytofluorometry and cytology. The DNA content of dissociated testicular cells of rats 6--30 days old, killed at daily intervals, was analysed by flow cytofluorometry using propidium iodide as a DNA-specific and quantitative fluorochrome. Testicular cells of a 6-day-old rat showed one peak of fluorescence. A second peak, at twice the modal channel number, appeared in testicular cells of 9-day-old animals. The number of cells under this peak increased progressively with age. A third peak, at half the channel number of the original one, appeared at 20 days and accounted for an increasing proportion of cells in testes taken from older rats. Cytological examination of the testicular tissue used for flow cytofluorometric analysis showed that preleptotene spermatocytes first appeared at 8 days after birth. Spermatids were first observed cytologically at 20 days after birth. The close temporal appearance of the fluorescence peaks with that of spermatocytes and spermatids, and the close association of the frequency of diploid and tetraploid cells as derived by flow cytofluorometry and cytology, indicated that the fluorescence peaks correspond--in order of increasing fluorescence--to spermatids, spermatogonia and somatic cells, and to spermatocytes. This conclusion was re-examined by analysing the ploidy levels of testicular cells of hypophysectomized or estradiol-treated by flow cytofluorodmetry. There was a loss of the haploid and tetraploid peaks subsequent to hypophysectomy. Estradiol dipropionate-treated rats, given weekly injections starting at 7 days of age, showed no appearance of the haploid peak and the regression of the tetraploid peak after an initial and transitory appearance. These results indicate that changes in ploidy levels that accompany the progression of meiosis in the testis were reflected in the sequential appearance of three fluorescence peaks as detected by flow cytofluorometry. The close correlation between the frequency of cell types as obtained by cytology and flow cytofluorometry indicates that the latter is a sensitive method for studying selected aspects of spermatogenesis in dissociated testicular cells.


Uteroglobin-like antigen in the pulmonary epithelium and secretion of the lung. The immunological identity of a uteroglobin-like protein, occurring in respiratory tract secretions and tissue, with uteroglobin from rabbit endometrial secretion is demonstrated. A uteroglobin-like antigen has been localized in bronchial epithelial cells and in bronchioles by immunofluorescence. This secretory protein is, in contrast to the authentic uteroglobin, hormone-independent, as far as estrogens and progesterone are concerned. The possible significance of comparative studies on uteroglobin and the uteroglobin-like antigen is discussed, taking into account cytological, endocrinological, and molecularbiological aspects.


Ultrastructure of rat pituitary LH gonadotrophs in relation to serum and pituitary LH levels following repeated LH-RH stimulation. The effects of single and repeated LH-RH injections at 120 min intervals on female rat LH gonadotrophs and on pituitary and serum LH levels were investigated using electronmicroscopy and radioimmunoassay. A temporary stimulation of granule release, of protein and new granule synthesis and of the accumulation of lysosomal structures was found in LH cells after the first LH-RH injection. The temporary stimulations were massively enhanced after the second injection. These consecutive yet in their time-sequence overlapping processes account for the initial depletion of secretory granule content (3--15 min after LH-RH injection), for the subsequent regranulation and accumulation of granules above control levels (60--120 min after injection) and also for the reduction in the number of granules to control levels (150 min after LH-RH injection and thereafter). Increased polymorphic lysosomal structures are believed to be responsible for this reduction of excess granules. The amount of assayable pituitary and serum LH generally corresponds with the morphological changes observed in LH-gonadotrophs, thus further substantiating the above observations. A schema which summarizes the observed morphological and hormonal changes in their time-sequence in response to LH-RH stimulation depicts the short-term regulation of secretory processes in female gonadotrophs.


Identification of the LH and TSH-secreting cells in the pituitary gland of the rhesus monkey. Pituitary glands from juvenile (pre-pubertal) and adult male and female rhesus monkeys were examined following immunocytochemical staining with antisera to the beta subunits of ovine luteinizing hormone (LHbeta) and of human thyroid stimulating hormone (TSHbeta). The LHbeta antiserum reacts with a cell that is PAS-positive, occurs singly and is randomly distributed throughout the pars distalis. The diameter of these cells is approximately 11.5 micrometer. They do not seem to vary in number in either juveniles (pre-pubertals) or adults, or in males or females. There appears to be fewer LH cells in the pituitary glands of pregnant and lactating females. In addition to staining cells in the pars distalis, the antiserum also reacts with a population of cells located in the pars tuberalis. The cells that stain with the anti-TSHbeta serum are confined primarily to the pars distalis. They are approximately 15.8 micrometer in diameter and are generally found in groups or clusters located in the anterior and medical regions of the gland. The TSH cells vary in number from one animal to another; however, this variability is unrelated to the age or the sex of the animals. No demonstrable changes occur in the number of TSH cells during pregnancy or lactation.


Immunocytochemical study of the neurohypophysial hormone producing system of the lungfish, Protopterus aethiopicus. Using the unlabeled antibody peroxidase-antiperoxidase complex (PAP) technique at the light microscopic level, it has been shown that, in the dipnoan preoptico-hypophysial neurosecretory system, vasotocin and mesotocin are synthesized in separate neurons. In the preoptic nuclei, the perikarya of these two types of neurosecretory neurons are not located preferentially. The two types of neurosecretory perikarya give rise to separate vasotocinergic and mesotocinergic axons, respectively. The dipnoan median eminence and neural lobe contain separate vasotocinergic and mesotocinergic nerve fibres, the general distribution of which is described. In the pars distalis and the pars intermedia of the hypophysis, neurohypophysial hormone-containing nerve fibres have not been found.


The ultrastructure of pinealocytes in the golden mole (Amblysomus hottentotus) with special reference to the granular vesicles. The ultrastructure of the pinealocytes of the golden mole (Amblysomus hottentotus), a blind subterranean mammal living in complete darkness, was examined and compared with that of pinealocytes of other mammals. On the basis of the presence of granular vesicles and glycogen granules, only one population of pinealocytes was found. Large mitochondria, ribosomes, lipid inclusions and scarce lysosomes were observed in the perikaryon. Numerous glycogen granules, often forming typical accumulations, were frequently found to be associated with typical vacuoles. An extraordinary large number of granular vesicles is characteristic of the golden mole pinealocyte. Phenomena of excretion of compound(s) stored in the granular vesicles were identified. Morphologically this is characterized by dissociation of its granular core in situ while its limiting membrane is preserved. The physiological significance of the secretory process characterized by the formation of these granular vesicles is discussed.


A Golgi study of the hypothalamus of Actinopterygii. II. The posterior hypothalamus. The posterior hypothalami of the polypteriform, Calamoichthys, and of the teleost, Anguilla, were studied by means of the Golgi technique. In Calamoichthys, the lateral lobes are not developed and the median lobe is simple. In Anguilla, the median (tuberal) lobe shows lophodendritic, CSF-contacting cells and horizontal cells in the periventricular grey and some reticular elements directed toward the cell-poor lateral areas. In the lateral lobes the periventricular grey is formed by multipolar neurons and a diffuse population of multipolar cells of uncertain identity. The nucleus diffusus lobi lateralis is formed by scarce multipolar neurons, often placed next to the external surface of the brain. The organization of the lateral lobes in Actinopterygii is reminiscent of highly developed integrative regions.


Intercellular attachments between calcified collagenous tissue forming cells in the rat. Osteoblasts of the young rat cranium, and cementoblasts and odontoblasts of young rat molars were prepared by ethanol freeze-fracture prior to critical point drying for scanning electron microscopy (SEM) as well as conventional transmission electron microscopy (TEM) techniques. Critical point drying causes shrinkage which separates the lateral intercellular contacts between neighbours in the same sheet in the case of cementoblasts and osteoblasts, but not those between odontoblasts. These differences are considered to be of functional significance and need to be taken into consideration when formulating theories of calcium influx into the mineralizable matrix of the respective tissues.


A stereological ultrastructural study of peritoneal macrophages from germ-free and conventionally-reared mice. The reported work is the first direct ultrastructural comparison of resident peritoneal macrophages from germ-free and conventional animals. Three groups of mice were studied: germ-free (GF), conventionally-reared under isolation conditions (IC), and conventionally-reared in an open environment (OC). The macrophages from the three groups of animals are closely similar morphologically. Particularly noteworthy are the electron-dense, lysosome-like granules which are numerous in the macrophages of germ-free mice and which provide a structural foundation for the presumed microbicidal capability of the phagocytes. Morphometric estimates showed that the "average macrophage" from GF mice is smaller and possesses a smaller, rounder nucleus, a smaller volume fraction of mitochondria and more lysosome-like granules per unit of cytoplasmic volume than the "average macrophage" from conventional mice. Moreover, granules and mitochondria are smaller, on average in the GF phagocytes than in macrophages from conventional mice. The results suggest that peritoneal macrophages from the germ-free mouse represent, more truly than those from the conventional mouse, the nature of the fully differentiated but as yet unstimulated mononuclear phagocyte.


Immunofluorescent staining of keratin fibers in cultured cells. Antibody prepared against a group of keratins purified from human stratum corneum was used to identify cells containing keratins by immunofluorescence. In sectioned tissue and in culture, keratinocytes of skin and other stratified squamous epithelia-whether human, rabbit of mouse-stained strongly, indicating homologous amino acid sequences in the keratins of these species. In all cases, the antibody revealed a dense cytoplasmic network of discrete fibers probably consisting of aggregated (tono-) filaments. The pattern of staining was not affected by cytochalasin B or colcemid. No keratins were detected in cultured cells of mesenchymal origin (3T3, NIL, BHK, human diploid fibroblasts) or in connective tissues, indicating that the 100 A filaments of fibroblasts are not related to the keratins. Keratinocytes at all stages of differentiation, including basal cells, stained brightly and therefore contained abundant keratins.


Stereo immunofluorescence microscopy: I. Three-dimensional arrangement of microfilaments, microtubules and tonofilaments. An easy manipulation of the commercial fluorescence microscope allows stereo pairs of pictures to be taken, which when examined with a stereo viewer, give a strong three-dimensional impression. The procedure is described in detail. Its use allows the documentation by immunofluorescence microscopy of the three-dimensional display and organization of microfilament bundles, microtubules and tonofilaments in some well characterized tissue culture cell lines.