A sample preparation for automated cervical cancer screening. A procedure for cluster disruption, cell dispersal and the production of a monolayer of cells from fluid suspensions for automated cell-scanning machines in an interactive system has been developed. The combination of 0.1 per cent dithiothreitol and 40 per cent alcohol in a balanced salt solution provides a self-limiting form of mucolysis which is completed by a controlled syringing regime to produce an optimally dispersed cell sample without the production of cell debris from the more effete cells. The induction of a positive charge on a microscope slide to attract the negatively charged cells has been achieved by coating the slides with the cationic polymer, polylysine. The cells appear to be sufficiently well attached to permit wet fixation and subsequent stain processing without loss. Quantitative data is given to demonstrate the superior cell presentation that is achieved by this new technique for automated scanning.


Glucose and cation transport in rat jejunum, ileum and colon in vivo: effects of anionic and nonionic surfactants, and of desoxycholate. Osmotically balanced solutions of glucose (0.5--300 mM) and sodium chloride, containing surfactants, were instilled into the small or large intestine of anaesthetized rats. Net absorption or secretion of glucose, sodium and potassium was studied. The surfactants tested were dodecylsulphate (3.4--17 mM), dioctylsulphosuccinate (1.8--11 m/M), Lubrol WX (0.1--5%), Triton X 100 (0.25%) and desoxycholate (2.5 mM). Qualitatively, the results were similar to those obtained previously with cationic compounds, suggesting a common mode of action for all surfactants studied. 17 mM dodecylsulphate seemed to abolish completely physiological glucose transport in the jejunum and ileum. At a lower concentration, and witth the other surfactants, normal glucose transport was affected to an intermediate extent.


Primary culture of adult rat liver cells. II. Cytological and biochemical properties of primary cultured cells. Primary mass culture of isolated cells from adult rat livers by the trypsin-perfusion method uas carried out to investigate cytological and biochemical properties of primary cultured cells. Two main types of cells were found in the course of primary culture of isolated hepatic cells. One was a group of polygonal cells with granular cytoplasm, which appeared to arrange themselves in cords and clusters 3 to 4 days postinoculation and then gradually decreased in number. These cells seemed to be associated with expression of liver specific functions such as higher tyrosine aminotransferase levels and transient increases in albumin production. Another type of cells was relatively small in size and had clear cytoplasm, which grew out rapidly in place of the polygonal cells and proliferated also in subculture. alpha-Fetoprotein was detected in the cultured media of the proliferating clear cells. These results suggest that the polygonal cells and the clear cells may correspond to the mature hepatocytes and the hepatocytic stem cells in vivo, respectively.


Acetic orcein staining of prefixed tissue sections. Acetic orcein stains formol- and Carnoy-fixed tissues, coloring mast cells, nuclei, basophilic cytoplasm, cerebral corpora amylacea, and cartilage strongly; keratin and erythrocytes moderately; muscle and collagen weakly. Guinea pig Brunner gland and rat colonic goblet cell mucins did not stain. The red nuclear stain contrasts well with the Prussian blue reaction of hemosiderin and the ferric ferricyanide (Turnbull's blue) reaction of enterochromaffin. A weak (0.01%) fast-green FCF stain changes collagen and sometimes smooth muscle to green, without impairing nucleic acid or mast cell staining. Picroindigocarmine gives blue collagen, yellow muscle, and red elastin, nucleic acids and mast cells. Picro-methyl blue tends to override the red nuclear stain. Carnoy fixation is somewhat better for nuclei, formol for basophil cytoplasms.


Staining of PO2 measuring points demonstrated for the rat brain cortex. A combined physiological and histological technique was developed in order to determine the exact position of the tip of a microelectrode used for PO2 measurements within the rat brain cortex. Using this method it was possible both to identify the tissue layer where PO2 measurements had been performed as well as obtain information about the blood capillaries within the surrounding tissue. After the PO2 had been measured small quantities of dye (Alcian Blue 8 GX) were injected through a micropipette attached to the PO2 microelectrode (tip-tip distance 2-5 micron). Subsequently the tissue was fixed by perfusion, 5 micron sections were cut and stained with Cresylviolet. The points where measurements had been made could be seen under a light microscope as green coloured spots of about 100 micron in diameter. The capillary pattern could be demonstrated by silver nitrate perfusion. Information about the capillaries, such as length, distance and the gas exchange surface were obtained via computer analysis of a television image. This method was used to analyse the O2 supply of the optical cortex of the rat. Correlation of the local PO2 distribution with the capillary data revealed differences between the archi- and the neocortex of the rat brain.


Amenorrhea and edema.

Amenorrhea and edema. In a 22 year old woman extensive edema developed during evaluation for amenorrhea. It was learned then that she was consuming escessive amounts of a laxative daily. Balance studies were performed which demonstrated that she excreted large amounts of sodium in her liquid stools while taking the dose of laxative she had been using. The presence of increased plasma renin activity and increased urinary aldosterone suggest that in this patient the occurrence of the edema after she discontinued use of the laxative was due to secondary aldosteronism presumably caused by plasma volume depletion.


Congenital corneal anesthesia.

Congenital corneal anesthesia. A 6-year-old boy with a visual acuity of 6/30 (20/100) in each eye had a diffuse corneal epithelial disruption that was most severe in the interpalpebral area. Corneal anesthesia was demonstrated with the Cochet-Bonnet aesthesiometer. Artificial tears, lubricant ointments, and taping of the eyelids were unsuccessful in ameliorating the problem. The use of therapeutic soft contact lenses provided visual acuity of R.E.: 6/12 (20/40), and L.E.: 6/6 (20/20), and arrested the epithelial breakdown. The contact lenses were discontinued after one year, and the corneal epithelium remained intact. Once the cycle of recurrent epithelial breakdown in congenital corneal anesthesia is interrupted for a prolonged period of time, one can expect a continued remission when therapeutic measures are discontinued. With early recognition and treatment of this disorder, corneal scarring can be avoided.


The inhibitory action of lidocaine in anaphylaxis. The action of lidocaine, a local anesthetic, was investigated during anaphylaxis in guinea pigs after passive sensitization in vitro of lung tissue and trachealis muscle. Pretreatment of the trachealis muscle with 8.54 mM lidocaine resulted in the total inhibition of anaphylactic isometric tension. Full reversal of anaphylactic-induced contractures was rapidly achieved with concentrations of 4.27 mM lidocaine. Release of histamine from both lung tissue and trachealis muscle was inhibited by 73 to 82 per cent, respectively, over concentration ranges of 2.13 to 8.54 mM lidocaine. A bimodal effect on sensitized tissues was noted, with lidocaine causing a slight release of histamine in the trachealis muscle of 1.6 per cent at a concentration of 8.54 mM. Lidocaine did not impair the initial passive sensitization process, nor did it appear to elute antibody once it was cell bound. The dual inhibitory effect on mast-cell release of mediators and on muscle contraction by lidocaine may be related in part to common processes involving the binding or flux of calcium.


Epidemiological features of the mycotoxicoses. Although mycotoxicoses attract more and more attention, the epidemiology of these diseases is still not adequately studied. The author devotes his study to certain diseases. In addition to the diseases whose mycotoxic nature is in no doubt, he conditionally includes certain diseases whose mycotoxic causation needs further proof. The author has assigned stachybotryotoxicosis and alimentary toxic aleukia or fusariotoxicosis to the first category and endemic nephropathy and primary liver-cell carcinoma to the second. His intention in adopting this approach is to establish the common features and differences in their epidemiological manifestations. Epidemiological studies not only help to define the incidence and prevalence of these disease but also make an important contribution to the determination of their nature. When stachybotryotoxicosis--the "unknown horse disease"--was being studied in the mid-thirties, in the Ukraine, the research workers headed by V. G. DROBOTKO, noticed that horses belonging to the Army remained unaffected while in the same localities those belonging to the collective farms were stricken by this "unknown disease". This epidemiological phenomenon suggested the hypothesis that the unknown diseases must be due to fodder, which was different for horses belonging to the Army and those belonging to the collective farms. This led members of the team to investigate the mycoflora of the fodder. The collective farmers themselves had noted the link between the disease and the feeding of poor quality musty straw to horses. For that reason in every collective farm where such cases occurred samples of the available fodder were studied. The numerous investigations remained fruitless for a long time until P. D. JATEL discovered straw which had been fed to horses before an outbreak of the disease. This straw contained a blackened layer copiously overgrown with the fungus Stachybotrys alternans. Feeding of fodder infected with that fungus to experimental horses reproduced the pattern of the "unknown disease", which thereafter began to be known as "stachybotryotoxicosis". When human stachybotryotoxocosis was studied, all the clues in the epidemiological investigation pointed to barley straw as a source of the disease. F. A. LINNIK (1938) noted that immediately before falling sick patients had been in close contact with musty straw. Among collective farmers the disease had a sudden onset and was mainly confined to men. No one contracted the disease who had not something to do with this musty straw. The members of the families of the patients also remained in good health. The disease bore a familial character when farmers had used the straw as litter on their private holdings. In their study of the nature of alimentary toxic aleukia, S. G. MIRONOV and M. K...


Failure of fenoprofen to affect the corneal immunologic reaction. In rabbits whose corneas had been subjected with bovine gamma globulin, fenoprofen administered subconjunctivally failed to alter the animals' inflammatory or immunologic responses.


[Gm(28), a new allotypic marker on human IgG3: peculiar interest of its study within Negroid populations (author's transl)]. A new allotype in the Gm system (Gm(28)) was described and studied. Among Caucasoids and Mongoloids, it was found with rare exceptions, in samples containing Gm(21), segregating with Gm1,17,21 and Gm1,2,17,21 haplotypes. This new antigenic determinant was found to be of particular interest among Negroid populations in which its frequency is variable. It can be detected with all--common or uncommon--haplotypes. The degree to which it is associated with some haplotypes in different populations may prove useful in the characterization of Negroid populations. This new allotypic determinant is located on the CH3 homology region of IgG3 subclass.


Congenital sensory neuropathy with selective loss of small myelinated fibers. The results of clinical, electrophysiological, autonomic, and histopathological studies in a 6-year-old patient with congenital sensory neuropathy are presented. Selective loss of pain perception affecting the extremities was noted from infancy. Nerve conduction studies were normal. Small myelinated fibers were selectively reduced in the sural nerve, and unmyelinated fibers were normal.


Glycoproteins in disease.

Glycoproteins in disease. The class of carbohydrate macromolecules known as glycoproteins exhibits great structural diversity. Their key role in the function of biological systems includes determination of cell membrane structure and function as well as antigenic determinants. In the recent decade, the detection of inborn errors of metabolism of these macromolecules as well as changes in their composition following mutagenesis and oncogenesis have highlighted the key role they play in the biology of man. The application of analytic techniques for their quantitation and structural characterization has resulted in their increasing utility in diagnostic medicine. They may be used not only as markers of active and inactive disease, but they also have become important in the diagnosis of congenital defects and tumors.


The structure of human immunoglobulins.

The structure of human immunoglobulins. General structural features of human Ig proteins are reviewed. The basic four-chain polypeptide structure is described with particular emphasis on IgG as a model for all Ig classes. The dual biologic role of antibody with antigen-binding and the Fc-associated class-specific effector functions is discussed. Certain aspects of Ig structure are highlighted in order to facilitate understanding of symposium topics related to Ig structure and function. Variable (V) region Ig structure, hypervariable segments in the V-regions, idiotypic determinants, and clonal switch mechanisms which permit the synthesis of IgM, IgG and IgA are presented in greater detail. Summary comments include a brief discussion of the genetic control of immunoglobulin synthesis.


African trials in chemotherapy of gastrointestinal cancer. Bleomycin either alone or in combination with radiotherapy was used in the treatment of 24 patients with non-resectable cancer of the oesophagus. Improvement in performance (swallowing) status was observed in 75% of patients, although only 19% were known to be alive after 1 year of follow-up. Patients with liver cancer were treated with adriamycin. The response rate was dose dependent and was 40% at best. Combination of adriamycin with other drugs did not improve the response rate. Intra-arterial adriamycin appears more efficacious and gave a response rate of 60%.


Management of malignant esophagorespiratory fistula. Esophagorespiratory communication developed in 46 patients among 570 with esophageal cancer. Therapy was basically palliative and aimed at mechanical interruption of the fistula, restoration of esophageal continuity, and avoidance of external tubes and appliances. Supportive therapy, gastrostomy, tracheostomy, and esophageal exclusion and diversion procedures resulted in little prolongation of life and poor palliation of the patient. Permanent endoesophageal intubation with tubes of the Celestin variety resulted in best palliation with minimal operative risk for most terminal patients. Colon bypass and occasional resection can accomplish the same goal and possibly provide long-term survival in good-risk, young patients with small tumors.


Burn unit management of toxic epidermal necrolysis. Toxic epidermal necrolysis is the name given to a group of dermatologic disorders characterized by a separation of epidermis and dermis with a subsequent skin slough. The denuded areas have the appearance of a second-degree burn. The complications of infection, negative nitrogen balance, severe pain, and emotional instability are identical to those seen in the patient with major burns. There are difficulties in patient management and advantages in burn unit care. As with the major burn, care of the patient with skin loss from toxic epidermal necrolysis is extremely complex, requiring the expertise of a burn team along with that of the dermatologist.


Serum biochemical markers in lung cancer. The prevalence of elevated serum levels of 5 potential tumour-associated antigens was determined in patients with lung cancer sampled at the time of initial presentation, using age- and sex-matched patients with benign lung disease as controls. Elevated levels (greater than upper 95th centile of controls) were found as follows: carcinoembryonic antigen (CEA), 17%; pregnancy-associated alpha-macroglobulin (PAM), 16%; casein 14%; human chorionic gonadotrophin (HCG) 6%; alpha-foetoprotein (AFP), 1.5%. The prevalence of elevated CEA levels (but not other markers) was higher in patients with evidence of extra-thoracic tumour spread (23%) mainly due to anaplastic tumours and adenocarcinomas. A degree of concordance of elevated marker levels occurred with CEA, HCG, casein and AFP, but there was a striking discordance of elevated CEA and PAM levels. Simultaneous assays of CEA and PAM will detect the majority of patients with elevations of any of the markers studied, and are likely to be the most useful biochemical markers in following the response of lung tumours to therapy.


Isolation of human tumour-specific antigens associated with beta2 microglobulin. In the present study the tube LAI assay was used to monitor the isolation of the TSA of 4 different types of human cancers. Each tumour antigen was found to be specific for tumours arising in the organ from which the TSA was initially derived and which were histopathologically similar. Immunochemical studies revealed that these molecules co-isolate with normal human HLA antigens and are associated with beta2m. On Sephadex G-150, the majority of the papain-solubilized tumour antigen eluted in the mol. wt range 70,000-150,000. Analysis of this material by SDS-PAGE and 6M guanidine-HC1 column chromatography indicated that the material is composed of smaller subunits with prominent peaks at approximately 40,000, 25,000 and 12,000 mol. wt. Immunoadsorbent affinity chromatography of the solubilized tumour-membrane constituents on AH-Sepharose-linked horse anti-human-beta2m indicated that the tumour antigens, like HLA molecules, contain a beta2m subunit. The specificity of binding of TSA to the immunoadsorbent columns and the immunologically specific abrogation of LAI reactivity were clearly shown. The present study, therefore, indicates that by the isolation of beta2m, human tumour antigens can also be isolated, since human tumour antigens are associated with beta2m. Whether human TSAs may perhaps be modified histocompatibility antigens remains to be answered. Although the change upon malignant transformation in the pattern of the cell-surface proteins expressing the TSA determinant remains obscure, it would appear that for tumours arising within a given organ, a consistent alteration of cell-surface proteins occurs.


Cellular immunity to encephalitogenic factor as measured by macrophage migration inhibition during tumour induction and growth. Spleen-cell sensitivity to encephalitogenic factor (EF) was measured with the macrophage migration inhibition (MMI) test over a period of time in hamsters inoculated with SV40-transformed tumour cells and in rats treated with 4-dimethylamino-3'-methylazobenzene.Spleen cells from hamsters receiving 10 or 10(3) SV40 tumour cells gave inhibition of macrophage migration with EF at a significance level of P<0·05 21 days after implantation. Spleen cells from animals receiving 10(5) tumour cells gave inhibition at a significance level of P<0·001 after the same interval.Spleen-cell sensitivity to EF, and the abrogation of this sensitivity by serum, was investigated over a period of time in rats undergoing hepatocarcinogenesis. Sensitivity to EF was seen in 2/10 animals (20%) with minimal lesions of the liver, in 2/16 animals (12%) with proliferative changes and/or cholangiofibrosis, in 7/15 animals (46%) with dysplastic lesions of portal-tract epithelial cells and in all 5 animals with cholangiocarcinoma. None of a control group of 10 animals showed any response to EF. Autologous serum abrogated the spleen-cell response to EF in one sensitized animal with proliferative changes and cholangiofibrosis, in all 7 sensitized animals with dysplastic hepatic lesions and in 4/5 sensitized animals with cholangiocarcinoma. Autologous serum had no effect on macrophage migration in the 10 control animals.These findings indicate that a progressive increase in sensitization to EF occurs during carcinogenesis and is evident at the point of preneoplastic dysplasia. This has an obviously important bearing on the clinical use of such tests.


Bleomycin-specific fragmentation of double-stranded DNA. Brief exposure of covalently closed circular duplex PM2 DNA to low concentrations of the clinical bleomycin mixture (Blenoxane) resulted in specific fragmentation of the genome that does not depend on the presence of superhelical turns. The double-strand breaks are in fact produced at several discrete sites on the PM2 genome but frequently occurring near the HpaII restriction endonuclease cleavage site. Initial rates of formation of nicked circular and linear duplex PM2 DNAs are reduced to different extents as the ionic strength of the reaction is increased. Increasing ionic strength is most effective in reducing the initial rate and overall yield of apparent double-strand scissions compared with single-strand scissions in the bleomycin-treated PM2 DNA.


Biochemical and immunochemical characterization of hexosaminidase P. Hexasaminidase P, the main isozyme of hexosaminidase in pregnancy serum, was isolated and purified 600--700-fold by a two-step purification procedure--affinity chromatography on Sepharose-bound epsilon-aminocaproyl-N-acetylglucosylamine, followed by ion-exchange chromatography on DEAE-cellulose. The purified enzyme was subjected to biochemical and immunochemical analysis. Its catalytic property, namely, kinetic behavior, is similar to that of the major isozymes of hexosaminidase, A and B. However, it differs from these isozymes in its electrophoretic mobility and in its apparent molecular weight which is around 150 000 compared with 100 000 of the A and B isozymes. Immunochemical analysis indicates that the P isozymes is antigenically cross-reactive with both A and B isozymes, but it does not contain the A-specific antigenic determinants, and exhibits identical antigenic specificity to hexasaminidase B. Two possible structures are suggested that are compatible with the experimental data: (a) a hexosaminidase B like structure with higher extent of glycosylation; (b) a hexameter of beta chain, possibly arranged as three beta2 subunits.


Complete covalent structure of human beta-thromboglobulin. The complete primary structure of the platelet-specific protein human beta-thromboglobulin has been determined. beta-Thromboglobulin consists of identical subunits of 81 amino acids, each with a molecular weight of 8851. The amino acid sequence of the beta-thromboglobulin subunit is: Gly-Lys-Glu-Glu-Ser-Leu-Asp-Ser-Asp-Leu-Tyr-Ala-Glu-Leu-Arg-Cys-Met-Cys-Ile-Lys-Thr-Thr-Ser-Gly-Ile-His-Pro-Lys-Asn-Ile-Gln-Ser-Leu-Glu-Val-Ile-Gly-Lys-Gly-Thr-His-Cys-Asn-Gln-Val-Glu-Val-Ile-Ala-Thr-Leu-Lys-Asp-Gly-Arg-Lys-Ile-Cys-Leu-Asp-Pro-Asp-Ala-Pro-Arg-Ile-Lys-Lys-Ile-Val-Gln-Lys-Lys-Leu-Ala-Gly-Asp-Glu-Ser-Ala-Asp. Disulfide bridge-18 to half-cystine-58. The amino acid sequence of beta-thromboglobulin shows a marked homology with that of platelet factor 4. When the sequences are aligned for maximum homology, 42 of the 81 residues of beta-thromboglobulin are identical with those of platelet factor 4, including the position of the four half-cystines.


The biosynthesis of oncovirus proteins.

The biosynthesis of oncovirus proteins. The patterns of oncovirus protein biosynthesis are essentially similar for avian and mammalian viruses. In each case the four major internal structural proteins are synthesized as a precursor polypeptide of about 75 000 daltons, the product of the gag gene. Translation occurs on genome-sized mRNA. This polyprotein is cleaved in a series of steps to give the mature proteins. The mechanism and localization of cleavage have not yet been clarified. Viral reverse transcriptase, the product of the pol gene, also is translated on genome-sized mRNA as a precursor, which is a "read-through" product of the neighbouring gag gene. The two major envelope proteins are translated as a glycosylated precursor of apparent molecular weight about 90 000 from the env gene located on a sub-genomic RNA species. The precursor is transported to the plasma membrane where it may mark the site of virus budding. It is cleaved in transport or on the membrane, but the resulting two mature envelope proteins remain tied by disulfide bonds. Sarc, the protein product of the src gene that is responsible for transformation, is translated from a different viral mRNA than the structural proteins. Sarc has not been definitively characterized in any system.


Kinetic analysis of the chlorophyll fluorescence inductions from chloroplasts blocked with 3-(3,4-dichlorophenyl)-1,1-dimethylurea. 1. The induction of Photosystem II chlorophyll fluorescence from chloroplasts blocked with 3-(3,4-dichlorophenyl)-1,1-dimethylurea and uncoupled with gramicidin has been measured. 2. In agreement with other authors it was found that the addition of cations to chloroplasts suspended in a low-cation medium not only stimulated the intensity of fluorescence but also changed the shape of the induction from being nearly exponential to being sigmoid. 3. A new theory of the photosynthetic unit of Photosystem II (Paillotin, G. (1976) J. Theor. Biol. 58, 237--252) was used to analyse the fluorescence inductions. 4. A comparison of the results of the Paillotin model with the experimental data suggests that excitation energy is not able to migrate between all the photosynthetic units of a photosynthetic domain. However, it is concluded that excitation energy may migrate from one photosynthetic unit to another, and that the energy migration is in competition with other processes leading to the decay of the excitation within Photosystem II. 5. It is suggested that the size of the "functional" photosynthetic unit, defined as the number of chlorophyll molecules that may communicate with a reaction centre, is variable.


Immunological properties of membrane-bound adenosine triphosphatase: immunological identification of rutamycin-sensitive F0.F1ATPase from Micrococcus luteus ATCC 4698 established by crossed immunoelectrophoresis. (1) F0.F1ATPase (EC 3.6.1.3) from Micrococcus luteus ATCC 4698 was solubilized from plasma membranes by the non-ionic detergent Triton X-100 in the presence of 0.05 M MgCl2. (2) The antibiotics rutamycin, Dio-9, quercetin, oligomycin, botrycidin, efrapeptin, leucinostatin, valinomycin, and venturicidin as well as N,N'-dicyclohexylcarbodiimide and dinitrophenol are potent inhibitors of F0.F1ATPase activity.(3) F0.F1ATPase activity is completely inhibited by anti-F1ATPase antibodies. The inhibition is non-competitive. (4) Crossed immunoelectrophoresis reveals a reaction of immunological identity of F0.F1ATPase and F1ATPase indicating that both enzymes have in common antigenic sites.


Molecular orbital studies on nucleoside analogs. II. Conformation of 6-azapyrimidine nucleosides. PCILO (Perturbative Configuration Interaction using Localised Orbitals) computations have been carried out for three 6-azapyrimidine nucleosides, 6-azauridine, 6-azacytidine and 6-azathymidine, for both C(2')-endo and C(3')-endo pucker of the sugar ring. The results indicate a syn (chiCN=180 degrees) conformation followed by chiCN=90 degrees and gg conformation for C(3')-endo 6-aza analogs as compareed to the anti (chiCN=0 degrees) and gg conformation preferred by the corresponding pyrimidine nucleosides. For C(2')-endo sugar geometry, 6-azauridine and 6-azacytidine prefer, respectively, chiCN=0 degrees (anti) and phi C(4')-C(5')=60 degrees C (gg) and chiCN-240 degrees (syn) and phi C(4')-C(5')=120 degrees. The corresponding nucleosides, uridine and cytidine, show a preference for syn (chiCN=240 degrees) and gg and anti(chiCN=0 degrees) and gg , respectively. The X-ray crystallographic conformations of 6-azauridine and 6-azacytidine have been attributed to intermolecular hydrogen bonding and crystal packing forces. The results of PMR, CD and ORD studies on 6-azauridine and 6-azacytidine in aqueous solutions are in agreement with the PCILO predictions.


Induction of four proteins in eukaryotic cells by kethoxal bis(thiosemicarbazone). Kethoxal bis(thiosemicarbazone) induces the synthesis of four proteins (100 000, 70 000, 35 000 and 25 000 daltons) in normal chick embryo cells. The 70 000 dalton species is produced at the fastest rate 2 hr after exposure to the compound. Pulse-chase experiments revealed neither precursors nor products of these proteins and both actinomycin and cycloheximide inhibited their synthesis. Neither of the two substituents of the inducer, kethoxal or thiosemicarbazide, were active. The four proteins were induced in several other species, but human cells produced only three proteins (100 000, 70 000 and a different 30 000 dalton form).


A rapid assay for neuraminidase. The detection of two differences in activity associated with virus transformation. Neuraminidase (acylneuraminyl hydrolase, EC 3.2.1.18) activity in fibroblast homogenates was measured by a rapid and simple assay with a synthetic substrate. The activity of neuraminidase in virus transformed hamster fibroblasts was increased over the normal counterpart. In addition, the differential activity seen using the synthetic substrate and fetuin made it possible to detect an enzyme activity hitherto not described. The advantages of this assay for metabolic screening are discussed.


A comparison of the polypeptide isoelectric points and antigenic determinant sites of the large subunit of fraction 1 protein from Lycopersicon esculentum, Nicotiana tabacum and Petunia hybrida. The large subunit of Fraction 1 protein from Lycopersicon esculentum, Nicotiana tabacum and Petunia hybrida has been examined by isoelectric focusing of the S-carboxymethylated polypeptides, and by double immunodiffusion with antiserum raised against Fraction 1 protein. The immunological results reveal heterogeneity in the large subunit primary structure not identified by isoelectric focusing. A variable phylogeny can be generated depending on whether serological or electrofocusing criteria are used.


[The antigen activity and structure of analogues and fragments of angiotensin, which contain enantiomeric forms of amino acids and aza-alpha'-homoamino acids]. The antigen activity of angiotensin, its fragments and analogues, which contain enantiomeric forms of amino acids and aza-alpha'-homoamino acids is studied by cross reactions with specific antibodies, obtained for angiotensin and its central tetrapeptide. It is found that the inclusion of additional NH-group between alpha carbon and carboxyl group of peptide chain, although in few cases radically changes spatial structure of compounds, does not deprivate their antigen activity. The replacement of NH-group to the C-end of the angiotensin molecule by affection the antigen determinant considerably decreases the antigen activity of the analogues. The important role in the determination of the antigen activity play the tyrosine residue and its specific orientation with respect to the antigen molecule. Low molecular weight fragments and analogues of the central part of angiotensin molecule, which have less "rigid" spatial structures, are capable to reorganize their spatial structure during interaction with antibodies.


Electrostatic calculations for an ion channel. II. Kinetic behavior of the gramicidin A channel. A theoretical model of the gramicidin A channel is presented and the kinetic behavior of the model is derived and compared with previous experimental results. The major assumption of the model is that the only interaction between ions in a multiply-occupied channel is electrostatic. The electrostatic calculations indicate in a multiply-occupied channel is electrostatic. The electrostatic calculations indicate that there will be potential wells at each end of the channel and, at high concentrations, that both wells can be occupied. The kinetics are based on two reaction steps: movement of the ion from the bulk solution to the well and movement between the two wells. The kinetics for this reaction rate approach are identical to those based on the Nernst-Planck equation in the limit where the movement between the two wells is rate limiting. The experimental results for sodium and potassium are consistent with a maximum of two ions per channel. To explain the thallium results it is necessary to allow three ions per channel. It is shown that this case is compatible with the electrostatic calculations if the presence of an anion is included. The theoretical kinetics are in reasonable quantitative agreement with the following experimental measurements: single channel conductance of sodium, potassium, and thallium; bi-ionic potential and permeability ratio between sodium-potassium and potassium-thallium; the limiting conductance of potassium and thallium at high applied voltages; current-voltage curves for sodium and potassium at low (but not high) concentrations; and the inhibition of sodium conductance by thallium. The results suggest that the potential well is located close to the channel mouth and that the conductance is partially limited by the rate going from the bulk solution to the well. For thallium, this entrance rate is probably diffusion limited.


Interactions in cation permeation through the gramicidin channel. Cs, Rb, K, Na, Li, Tl, H, and effects of anion binding. As a prototype for binding and interaction in biological Na and K channels, the single channel conductances for Li, Na, K, Rb, Cs, H, and Tl and the membrane potentials for Tl-K mixtures are characterized for gramicidin A over wider concentration rangers than previously and analyzed using an "equilibrium domain" model that assumes a central rate-determining barrier. Peculiarities in the conductance-concentration relationship for TlF, TlNO3, and TlAc suggest that anions bind to Tl-loaded channels, and the theory is extended to allow for this. For concreteness, the selectivity of cation permeation is characterized in terms of individual binding and rate constants of this model, with the conclusions that the strongest site binds Cs greater than Rb greater than K greater than Na greater than Li, while the next strongest binds Na greater than K greater than Li greater than Rb greater than Cs. However, because Schagina, Grinfeldt, and Lev's recent finding of single filing (personal communication) indicates that the channel sites in gramicidin cannot be at equilibrium with the solution, and work in progress with Hägglund and Enos (Biophys. J. 21:26a. [Abstr.]) indicates that the simplest model adequate to account for the observed concentration-dependences of flux-ratio, conductance, I--V characteristic, and permeability has three barriers and four sites, some implications of additional rate-determining barriers at the mouth of the channel are discussed. The results are summarized using phenomenological "experimental" parameters that provide a model-independent way to represent that data concisely and which can be interpreted physically in terms of any desired model.


[Localization of specific antigen in the organs of animals inoculated with different series of live measles vaccine]. Specific antigen was identified by the immunofluorescent test in the walls of the brain blood vessels, in the neurons, and in the glial cells of albino newborn mice and Syrian hamsters inoculated subcutaneously with 4 different batches of live measles vaccine. The pathomorphological test of the brain tissue revealed mainly vascular disturbances. The data obtained testify to the presence of residual neurotropism in the attenuated measles virus (strain "L-16").


[Decreased anaphylactogenic properties of disaggregated gamma-globulin]. Immunological properties of native, aggregated, and disaggregated (tolerogenic) human gamma-globulin were compared in experiments on mice and guinea pigs. Disaggregated preparation was shown to give diminished local and general hypersensitivity reaction in sensitized animals and had decreased ability to induce secondary humoral immune response.


[Comparative immunochemical and physicochemical characteristics of human placental chorionic alphal- and alpha2-microglobulins]. Immunochemical analysis showed chorionic alpha1-microglobulin to be immunologically different from chorionic alpha2-microglobulin. Some physico-chemical properties of these proteins were studied, and they were found to differ from one another by a number of parameters.


[Localization of albumin, transferrin and alpha-fetoprotein in normal and regenerating mouse liver]. An acetone-formol fixation technique with subsequent paraffin-embedding suitable for immunofluorescent study of different antigens, including serum proteins, is described. This technique was used for detection of albumin, transferrin, and alpha-fetoprotein distribution in the normal and regenerating liver of mice. Albumin and transferrin were always found together in the same hepatocytes, both under normal conditions and in regeneration. In the regenerating liver alpha-fetoprotein was encountered independently of the two other proteins, although it was revealed in the same zones. Only in the perinecrotic zone did each alpha-fetoprotein-positive hepatocyte contain albumin and transferrin.


Development of a practical oral dexamethasone premedication schedule leading to improved granulocyte yields with the continuous-flow centrifugal blood cell separator. Studies were conducted to improve the yield of granulocytes collected for transfusion from normal donors by means of the continuous-flow centrifugal blood cell separator. Nonleukapheresed donors were medicated with varying schedules of corticosteroids to learn the magnitude and duration of granulocytosis. Normal donors were medicated with varying schedules of corticosteroids prior to a 4-hr leukapheresis and the granulocyte yields determined. It was found that maximum yields (32.2 x 10(9) granulocytes) were obtained by use of dexamethasone given orally 12 and 3 hr prior to leukapheresis. There was a good correlation between the yields and the circulating granulocyte count at the start and during the procedure.


Factors influencing postoperative morbidity and mortality in patients treated with bleomycin. Patients treated with bleomycin are at risk of developing the acute adult respiratory distress syndrome post-operatively. In a prospective study of 12 patients who had received bleomycin preoperatively and were undergoing removal of retroperitoneal lymph nodes or pulmonary metastases several preventive factors were established. These were the use of low concentrations of inspired oxygen during operation and in the immediate postoperative period, careful monitoring of fluid replacement, and restriction of crystalloids in favour of colloids.


A quantitative electron microscopic study of synapse formation in dispersed cell cultures of rat cerebellum stained either by Os-UL or by E-PTA. Synapse formation was followed in dispersed cell cultures of rat cerebellums stained either by osmium-uranyl-lead (Os-UL) or by ethanolic phosphotungstic acid (E-PTA). The numerical densities of synapses stained by Os-UL were always significantly higher than those stained by E-PTA (from 3 to 35 days in vitro). This difference suggests that some portion of the populations of both immature and mature synapses was not stained by E-PTA. The width of the synaptic cleft (28.4nm) in synapses stained by E-PTA was only 9nm more than that of the cleft in synapses stained by Os-UL (19.4nm), suggesting that some portion of one or both of the synaptic membranes is stained by E-PTA. Analysis of data from 7 other staining procedures utilizing both ethanolic and aqueous solutions demonstrated that the differences in cleft width described above appear to be due to the various affinities of the stains for different portions of synaptic membranes, and do not represent differences due to shrinkage artifact. In examining the parameters of synaptic structure during development of the cultures, a statistically significant increase in both the height and width of the presynaptic dense projections was found. Changes in synaptic morphology during synaptogenesis in this culture system were similar to those reported for the cerebellar cortex in vivo.


Horseradish peroxidase labeling of cat tectotectal cells. By means of unilateral intracollicular injections of horseradish peroxidase (HRP), the cells of origin of the tectotectal projection in cats were detected in the opposite colliculus. The 9 injected animals were divided into two groups in accordance to the location of their injections along the collicular rostrocaudal axis. The majority of the labeled cells were obtained from two injections performed at the level of the commissure of the superior colliculus (231 cells) whereas 5 injections performed at lower levels led to labeling of 49 cells. In two animals, the injections did not cause labeling of cells, a fact that was attributed to the superficiality of the injections. Concerning the distribution of the labeled cells over the collicular layers and along the collicular rostrocaudal axis, there was no difference between the two groups of animals. Among the 280 labeled cells, 1 cell (0.35%) was found in stratum griseum superficiale, 83 cells (29.64%) in stratum opticum, 182 cells (65.00%) in stratum griseum intermediale and 14 cells (5.00%) in stratum griseum profundum. Ninety-two per cent of the cells were located in the rostral portion of the colliculus, in the region between the collicular rostral tip and the collicular rostral plane that corresponds to the vertical meridian representation. Eight per cent of the cells were concentrated in a zone comprising the lower half of stratum opticum and the upper half of stratum griseum intermediale, a location that corresponds to the transition between the visual-sensory and the visual-motor colliculus. On the basis of laminar segregation of collicular functions, the possibility that the tectotectal projection could be related to visual orienting, following and attentive collicular functions through the suppression of activity (a non-enhancement effect?) of homologous cells in the opposite colliculus is discussed.


Vestibular nucleus neurons relaying excitation from the anterior canal to the oculomotor nucleus. A morphological approach was undertaken to determine which vestibular nucleus neurons relay excitation from the anterior canal to the IIIrd nucleus. In anesthetized rabbits HRP was iontophoresed into the IIIrd nucleus and cells filled with HRP reaction product (positive cells) searched for within the vestibular nuclear complex. By lesioning the MLF or brachium conjunctivum immediately after iontophoresis it was demonstrated that positive cells in the dorsum of the superior vestibular nucleus are backfilled via their axons which ascend in the brachium conjunctivum. By contrast positive cells in the center of the superior nucleus are backfilled via their axons in the MLF. In electrophysiological experiments in the presence of a severed MLF the anterior canal was selectively stimulated for orthodromic, and the 3rd nucleus stimulated for antidromic, activation of vestibular nucleus neurons. Recording extracellularly with glass microelectrodes filled with fast green FCF the only cells both ortho- and antidromically activated were localized to the dorsum of the superior vestibular nucleus. It is concluded that cells dorsally located in the superior nucleus relay the disynaptic excitatory vestibulo-ocular reflex from the anterior canal to the contralateral 3rd nucleus via their axons which ascend in the brachium conjunctivum.


A sensitive method for histochemical demonstration of horseradish peroxidase in neurons following retrograde axonal transport. A study was made on the effects of various fixatives and some other histochemical parameters used in the procedure for demonstrating labeled neurons following retrograde axonal transport of horseradish peroxidase (HRP). The enzyme was injected into the tongue of adult mice and the results were obtained by counting labeled hypoglossal neurons following certain variations in the procedure. Paraformaldehyde in the fixative should be avoided since it reduces the number of labeled neurons as compared to glutaraldehyde in a concentration of 1.5-2.5% Fixation for about 4 h is recommended followed by a wash in 5% sucrose buffer overnight. Variables in the histochemical procedure were systematically studied in order to determine optimal pH, buffer type, buffer concentration and substrate concentration. The effect of using a "preincubation" in buffer containing only diaminobenzidine tetrahydrochloride (DAB) was also examined. These results were used to develop a modified histochemical procedure which produced a substantial increase in the number of detectable HRP-labeled neurons as compared to equivalent sections that were reacted in the incubation medium described by Graham and Karnovsky. The modified histochemical procedure involves incubation (no preincubation with DAB only) of sections in the dark for 30 min in a solution consisting of 10 ml cacodylate buffer (pH 5.1;0.1 M), 20 mg DAB and 0.1 ml of 1% hydrogen peroxide. The Kodak Wratten no. 46 filter is recommended for light-microscopical identification of labeled neurons since it is closely matched to the absorption spectrum of the DAB reaction product and consequently greatly increases the contrast of HRP-labeled neurons.


Radial glia in the human fetal cerebrum: a combined Golgi, immunofluorescent and electron microscopic study. Golgi techniques, immunofluorescence for glial fibrillary acidic (GFA) protein, and electron microscopy (EM) were used to determine the nature of radial glia in the cerebrum of human fetuses ranging from 7 to 20 weeks of ovulation age. Successful Golgi impregnation of radial fibers was achieved in fetuses 12 weeks of age and older. These fibers spanned the entire thickness of the hemisphere. At the pial surface many of them branched and terminated in pyramidal end feet expansions. Indirect immunofluorescent preparations utilizing antiserum to GFA protein, a protein specific for astrocytes, demonstrated numerous radially oriented nearly parallel fluorescent fibres between the ventricular zone and pia mater. GFA protein-positive fibers were demonstrated in all fetal specimens examined with this technique (10 weeks of age and older). Along the outer border of the marginal zone they formed a horizontal GFA protein-containing subpial membrane. By EM there were numerous linear electron lucent astrocytic processes containing 8-9 nm filaments and occasional glycogen granules at all levels of the cerebrum. They were interspersed among smaller and darker neuronal processes containing 20-25 nm neurotubules, and were demonstrable at all fetal ages between 7 and 18 weeks. They formed pericapillary investments and subpial terminal expansions closely abutting basal lamina of pia mater in every specimen examined. On the basis of these combined analyses, we conclude that radial glial fibers in early human fetal cerebrum represent processes of immature astrocytes. Although subsequently undergoing further maturation, radial glia already possess fundamental immunocytochemical and morphological characteristics indicative of astrocytic differentiation. A significant implication of our findings is that the development of astrocytes in the human fetal brain occurs much earlier than formerly believed.


Effect of antisera to beta and gamma goldfish brain proteins on the retention of a newly acquired behavior. The metabolism of 3 brain cytoplasmic proteins (alpha, beta, and gamma) increases markedly when goldfish acquire a new pattern of behavior. Antisera specific to beta and beta + gamma proteins were prepared and injected into the fourth ventricle of the brains of trained animals at 8 and 24 h after the initiation of training. When tested 3 days later, such goldfish (N = 98) could not recall the training; whereas trained goldfish (N = 97) receiving non-immunized rabbit serum had complete recall of the behavior. Also no amnesia was obtained in control experiments in which trained goldfish were injected with an antiserum to a neural surface membrane protein NS-6. The fact that antisera to beta + gamma had no toxic effects was demonstrated by injecting them prior to training; no effects on the rate of acquisition and recall of the behavior was found. The antisera to beta + gamma were effective in inhibiting recall of the training when they were injected any time between 3 h up to 48 h after training; no effect was obtained at 72 h post training. These results are consistent with the hypothesis that beta and gamma might have some functional role in the plasticity of the CNS.


Reversible reduction of tyrosine hydroxylase enzyme protein during the retrograde reaction in mesolimbic dopaminergic neurons. Changes in the activity and amount of the neurotransmitter synthesizing enzyme tyrosine hydroxylase (TH) were measured in dopaminergic (DA) neurons of the A10-mesolimbic system of the rat following electrolytic lesions of their axons. Unilateral hypothalamic lesions in close proximity to the cell bodies resulted, within 24--48 h, in a permanent anterograde reduction of TH to 10--20% of control in the ipsilateral olfactory tubercle and nucleus accumbens. The retrograde reaction in the A10-DA nerve cell bodies was characterized by an initial increase in TH activity to 133% by 24--48 h followed by a gradual and permanent fall to 50% of control by day 14 due to retrograde cell death of DA neurons. In contrast, lesions of DA axon terminals in the olfactory tubercle resulted in a reversible retrograde reduction of TH activity of the A10. The enzyme activity declined during the first 7 days to 70% of control and then gradually recovered, reaching control levels by 28 days after the operation. The reduction in TH activity in the A10 was demonstrated by immunochemical titration with a specific antibody to TH to be entirely due to reduced amounts of enzyme protein. We conclude that in mesolimbic DA neurons; (a) the anterograde reaction is characterized by a rapid and permanent decline of TH in degenerating terminals; (b) the retrograde reaction is dependent upon the proximity of the lesion to the nerve cell body, and (c) a reversible reduced accumulation of TH characterizes the retrograde reaction in response to lesions of distal axons. The reaction of DA neurons of the mesolimbic system to axonal injury is comparable to that of the nigrostriatal system.


Rh isoimmunization during pregnancy: antenatal prophylaxis. Of 3533 Rh-negative women who began a pregnancy without detectable Rh antibodies, 62 (1.8%) demonstrated evidence of Rh isoimmunization during pregnancy or within 3 days after delivery. All denied transfusions as well as abortions or previous pregnancies not followed by the administration of Rh immune globulin. Rh isoimmunization during pregnancy or within 3 days after delivery, which will not be prevented by the administration of Rh immune globulin after delivery, is the most important cause of residual Rh isoimmunization. A clinical trial of antenatal administration of Rh immune globulin, initially at 34 weeks's and subsequently at 28 and 34 weeks' gestation, in 1357 Rh-negative pregnant women who were delivered of Rh-positive babies, was effective in preventing the development of Rh isoimmunization during pregnancy or within 3 days after delivery. Antenatal prophylaxis  with Rh immune globulin will be necessary if the incidence of Rh isoimmunization is to be reduced to its lowest possible level. Antenatal prophylaxis at 28 weeks' gestation is now an insured service in Manitoba.


Antenatal prophylaxis of Rh isoimmunization: 28-weeks'-gestation service program. Two (0.18%) of 1086 Rh-negative primigravidas or multigravidas treated similarly in all previous pregnancies, who were given a single injection of Rh immune globulin (300 mug) at 28 weeks' gestation and subsequently were delivered of Rh-positive babies, had demonstrable Rh isoimmunization at the time of that injection and must be considered "logistic" failures of antenatal prophylaxis. The remaining 1084 (who were treated again after delivery) had no evidence of Rh isoimmunization at delivery and none of the 512 screened at 6 months after delivery appeared to be immunized. If the 28th-week injection had not been protective, one would have expected 14 of the 1084 to have been demonstrably Rh isoimmunized and evidence of Rh isoimmunization to have persisted in 6 of the 512 observed 6 months after delivery.Six of 719 Rh-negative multigravidas who had not received Rh immune globulin after previous pregnancies or had been treated only after delivery showed evidence of Rh isoimmunization despite a single injection of Rh immune globulin at 28 weeks in a subsequent pregnancy. In three of the six the cause was most likely "sensibilization" due to previous exposure to Rh-positive blood or an untreated Rh-positive pregnancy. in 3 of the remaining 716 (0.42%) there may have been true failure of antenatal Rh prophylaxis administered at the 28th week. One would have expected this figure to be 12 of 716 if antenatal Rh prophylaxis at 28 weeks' gestation were totally unsuccessful.It is concluded that a single intramuscular injection of Rh immune globulin, 300 mug, is 88% effective in preventing Rh isoimmunization during pregnancy in Rh-negative primigravidas and in multigravidas treated antenatally in all previous pregnancies, and is 75% effective in preventing Rh isoimmunization in Rh-negative multigravidas untreated during previous pregnancies. The majority of failures are due to Rh isoimmunization during pregnancy prior to antenatal prophylaxis at 28 weeks.


Treatment of advanced Hodgkin's disease with adriamycin, bleomycin, vinblastine, and imidazole carboxamide (ABVD) after failure of MOPP therapy. From June 19, 1975 to December 22, 1976, twenty-seven patients with advanced Hodgkin's disease who failed MOPP (nitrogen mustard, vincristine, procarbazine and prednisone) were treated with adriamycin, bleomycin, vinblastine, and imidazole carboxamide, (ABVD). Complete response (CR) was achieved in 22% of patients and partial response was achieved in 15%. No response was observed in 63% of patients. With a median duration of follow-up for CR patients of only 10.5 months, two of the six CR patients have already relapsed. In this series of patients ABVD was not an effective curative regimen for patients with Hodgkin's disease who have failed MOPP.


Radiation nephritis following combined abdominal radiation and chemotherapy (bleomycin-vinblastine). A 29-year-old man presented with acute glomerulonephritis five weeks following completion of combined chemotherapy (bleomycin-vinblastine) and abdominal radiation for testicular carcinoma. There was no evidence for a post-infectious cause or a systemic collagen disorder. The renal biopsy showed changes consistent with radiation nephritis. The combined radiation and chemotherapy may have, by additive or synergistic action, caused the early appearance of radiation nephritis.


A phase I study of active specific intralymphatic immunotherapy (ASILI). Twenty-one patients with advanced malignancies who had exhausted or refused conventional modalities of treatment were entered in a Phase I toxicology trial of active specific intralymphatic immunotherapy (ASILI). The patients were immunized with 1 X 10(7) to 1.2 X 10(8) viable autochthonous or allogeneic irradiated tumor cells intralymphatically each month and received no other antineoplastic treatment. To date, 274 intralymphatic injections have been performed and except for one case of bacterial lymphangitis, no adverse side effects have been observed. ASILI did not significantly alter peripheral blood lymphocyte counts, absolute E-rosette forming cell levels, or EA-rosette forming cell levels. PHA reactivity of peripheral blood lymphocytes increased slightly in all but one patient tested. Seven out of nine patients who had not had delayed hypersensitivity to recall antigens developed positive reactions following ASILI. Sixteen out of twenty patients tested also developed reactivity to their immunizing cells after treatment. Objective regression (greater than 50% reduction of tumor mass) was observed in five out of nineteen evaluable patients. Six patient showed stabilization of tumor growth and eight patients continued to progress under treatment.


Bone marrow acid phosphatase by radioimmunoassay. A double-antibody radioimmunoassay was developed and utilized to measure prostatic acid phosphatase in bone marrow aspirates. One hundred-eighteen patients with carcinoma of the prostate in various clinical stages, and fifty with benign prostatic hyperplasia were studied. In patients with carcinoma, levels of prostatic acid phosphatase in bone marrow aspirates were found to correlate well with increasing clinical stage of the disease. Determination of bone marrow prostatic acid phosphatase by radioimmunoassay may be a valuable adjunct to clinicopathologic staging of prostatic carcinoma.


Alpha-fetoprotein and human chorionic gonadotropin in a patient with a primary intracranial germ cell tumor. A 17 year old man with a primary intracranial germ cell tumor in the pituitary region is reported. Both serum alpha-fetoprotein (AFP) and serum human chorionic gonadotropin (hCG) were found elevated preoperatively. Normal levels of serum AFP and hCG were found after operation and radiotherapy. By indirect immunofluorescence staining of the tumor tissue a slight staining for AFP was localized to the endodermal vesicles, whereas a more intensive staining for hCG could be identified in the giant cells.


A comparison of acridine orange and Feulgen cytochemistry of human tumor cell nuclei. Specimens of cells derived from tumors of the human female genital tract plus normal cells as standards have been divided into aliquots and stained according to acridine orange or pararosanilin:Feulgen procedures. Acridine orange-stained cells were slit-scanned for 535 nm nuclear fluorescence; Feulgen-stained cells were comb-scanned for 580 nm nuclear absorbance. For each specimen examined, the tumor cell:normal cell ratio of mean nuclear fluorescence following acridine orange staining was greater than the tumor cell:normal cell ratio of mean nuclear absorbance following Feulgen staining. The tumor cell:normal cell ratio of mean nuclear fluorescence ranged from 2.3 for a nonkeratinizing squamous cell carcinoma to 3.9 for a keratinizing squamous cell carcinoma. The tumor cell:normal cell ratio of mean nuclear absorbance ranged from 1.4 for a mixed mesodermal sarcoma to 2.3 for a small cell squamous cell carcinoma. These results indicate that the elevated nuclear fluorescence intensity from acridine orange-stained tumor cells cannot be explained solely on the basis of elevated Feulgen:DNA content. An alternative hypothesis, consistent with these results, is that DNA is the principal binding substrate for intranuclear acridine orange and that the DNA of certain tumor cells is more accessible to acridine orange than is the DNA of normal cells.


Differences in nucleolar antigens of rat liver and Novikoff hepatoma ascites cells. Antisera to nucleoli of Novikoff hepatoma ascites and normal rat liver cells were produced in rabbits by injection of whole, isolated nucleoli. These antisera have been used to compare the nucleolar antigens that were partially fractionated by differential solubilization from nucleoli. Fourteen antigens were detected by these antisera; ten of these antigens were detected by both antisera. Ouchterlony double diffusion analysis of soluble extracts from normal rat liver and Novikoff hepatoma ascites nucleoli and fetal rat liver nuclei provided evidence for antigens found only in liver extracts, only in tumor extracts, or only in tumor and fetal extracts. Antisera preabsorbed to remove antibodies to common antigens of liver and tumor provided confirmatory evidence for one nucleolar antigen in liver that was not found in tumor or fetal rat liver, one antigen in tumor that was not found in adult or fetal rat liver, and three antigens in both tumor and fetal rat liver that were not found in adult rat liver. In addition, the antitumor nucleolar antiserum preabsorbed with liver nuclear extracts still produced positive nucleolar fluorescence in Novikoff hepatoma ascites cells but not in liver cells. Conversely, anti-liver nucleolar antiserum preabsorbed with tumor nucleolar extracts did not produce detectable tumor nucleolar fluorescence but did produce positive fluorescence in liver nucleoli.


Radioimmunoassays that demonstrate type-specific and group-specific antigenic reactivities for the major internal structural protein of murine mammary tumor viruses. The 28,000-dalton (p28) major structural polypeptide of the mouse mammary tumor virus (MMTV) was isolated and used to develop a highly sensitive and specific radioimmunoassay. Under conditions of limiting antibody in competitive binding assays, as little as 50 pg of purified p28, as well as disrupted MMTV virions and mammary tumor extracts, competed specifically with 125I-labeled MMTV p28. The p28 polypeptide was further shown to contain both group-specific and type-specific antigenic determinants, thus also allowing for further differentiation of various MMTV strains.


Comparative antitumor activity of actinomycin analogs in mice bearing Ridgway osteogenic sarcoma or P388 leukemia. Actinomycin (Act) analogs, differing in the chemical substitution(s) made at various positions in either their pentapeptide chain(s) or chromophore ring, were evaluated for their antitumor activity in mice bearing either Ridgway osteogenic sarcoma (ROS) or P388 leukemia. Of the analogs tested against advanced (2--3-g) ROS tumors, azetomicin I and Act III caused therapeutic responses which, although variable, were nevertheless indicative of antitumor activities greater than was found using Act D. Several other analogs, Act C2, 2-N-(gamma-hydroxypropyl)-Act D, Act X0delta, and azetomicin II, displayed antitumor activity in ROS-bearing mice which varied, in different experiments, from comparable to superior to that achieved using Act D. Additionally, Act Pip1beta and 3'-(4-cisCl-Pro)-Act were comparable to, and Act-2-hydroxy-C3 inferior to, Act D in activity against ROS. Both azetomicin I and II were as effective as Act D in mice bearing P388 leukemia. Moreover, a subline of P388 that is resistant to Act D was cross-resistant to both azetomicin I and II.


Analysis of a 5' leader sequence on murine leukemia virus 21S RNA: heteroduplex mapping with long reverse transcriptase products. The majority of the mRNA that specifies retrovirus glycoproteins is known to be derived from the 3' half of the genome. To examine whether the glycoprotein mRNA of murine leukemia viruses (MuLVs) might consist of portions derived from both the 5' and 3' ends of the viral genome, we performed hybridization with a 5'-specific probe and heteroduplex analysis with long reverse transcribed DNA. A 5' probe was made by purifying a discrete 50 nucleotide-long reverse transcript attached to its tRNA primer. This probe was found to hybridize to RNA of the size of glycoprotein mRNA--21S, poly(A)-containing RNA--indicating that the mRNA could have a 5' leader sequence. The 5'-specific sequences were studied by electron microscopic examination of hybrids between 21S RNA and the two longest discrete cDNA species synthesized in the endogenous reverse transcriptase reaction. One of these species, 8.8 kb long, is only made in the absence of actinomycin D, but it does not contain any self-complementary sequences, and therefore appears to be a complete transcript of the viral genome. The shorter of the two species, 8.2 kb long, is synthesized whether or not actinomycin D is present; it must terminate 500--600 nucleotides internal to the 5' end of the template RNA. The structures observed in heteroduplexes of 21S RNA and these DNAs indicated the presence of a leader sequence approximately 500 nucleotides long at the 5' end of the 21S RNA. Sequences comprising this leader segment in the 21S RNA mapped at the 5' end of the genome RNA; the rest of the 21S RNA consisted of sequences from the 3' portion of the genome. Analysis of heteroduplexes with 8.2 kb DNA suggested that actinomycin D could block the reverse transcription of most of the sequence in the genome RNA that appears as a leader in the 21S RNA.


Merocyanine 540 as a fluorescent probe of membranes: staining of electrically excitable cells. With the exception of certain blood cells considered in the accompanying paper (Valinsky, Easton and Reich, 1978), merocyanine 540 (MC 540), a fluorescent membrane probe, selectively strains the membranes of a wide variety of electrically excitable cells, but not those of nonexcitable cells. This reaction is Ca2+-dependent when staining is performed in buffered iso-osmotic sucrose, Ca2+-independent when staining proceeds at high ionic strength, inhibited by La3+ and sodium Suramin, enhanced by controlled, low level photosensitization of cell-associated dye and essentially irreversible. These characteristics of the staining reaction depend upon the maintenance of both cell viability and a normal unperturbed membrane structure. Although the mechanisms involved in the staining specificity remain unknown, observation of MC 540 partitioning between benzene and water in model reactions indicates that dye transport into hydrophobic solvents is accompanied by the formation of stoichiometric complexes with cations and phospholipids. These results may suggest the existence of specific, possibly phospholipid-rich membrane domains that mediate complex formation with MC 540 in excitable cells; comparable domains either would not exist, or would be inaccessible at the external surfaces of nonexcitable cells.


The visualization of fluorescent proteins in living cells by video intensification microscopy (VIM). A highly sensitive television camera (silicon intensifier target) has been combined with fluorescence microscopy to examine living cultured cells. This system is termed Video Intensification Microscopy (VIM). By using very small amounts of excitation light, one limits the damage to living cells from excessive illumination and is able to visualize fluorescence probes for periods up to 24 hr without bleaching. With VIM, the cellular uptake and fate of two rhodamine-labeled proteins, concanavalin A and alpha2 macroglobulin, have been followed for up to 24 hr. These proteins were first located in endocytic vesicles with a low phase density. Later, at 24 hr, alpha2 macroglobulin was located in phase-dense structures, probably secondary lysosomes. Both the fluorescent endocytic vesicles and lysosomes were observed to undergo saltatory motion. VIM combined with fluorescence promises to have a widespread application in the study of the behavior of living cells.


Merocyanine 540 as a fluorescent probe of membranes: selective staining of leukemic and immature hemopoietic cells. We have reported (Easton, Valinsky and Reich, 1978) that merocyanine 540 (MC 540) specifically stains a variety of living excitable cells, but not nonexcitable cells. This paper describes the exceptional permeability to MC 540 of leukemic leukocytes and immature hemopoietic precursor cells. We have used fluorescence microscopy and uptake of radioactive dye to study MC 540 staining of peripheral blood leukocytes from 80 leukemic and 34 normal individuals; leukemic leukocytes stain, whereas normal leukcytes do not. The leukocyte staining reaction differs from that previously described for excitable cells since it is independent of the ionic composition of the staining medium, kinetically complex, enhanced by light, enhanced by oxygen and essentially irreversible. Virtually all circulating nucleated cells from leukemic individuals are stained to approximately the same extent, and there is no qualitative or quantitative distinction between the various forms of leukemia. We have also found that MC 540 interacts with granulopoietic colony-forming cells (CFU-C) and with spleen colony-forming cells derived from mouse bone marrow (CFU-S). We cannot as yet identify a specific property of leukocyte plasma membranes that determines MC 540 permeability; since changes in MC 540 uptake appear to be correlated with cellular maturation during normal hemopoiesis, the retention of staining by leukemic cells, some of which appear morphologically normal, may indicate of failure in membrane maturation during leukemic blood cell development.


The HLA-dependent expression of testis- organizing H-Y antigen by human male cells. The proposal that the stable expression of organogenesis-directing plasma membrane antigens, such as testis-organizing H-Y antigen, requires beta2-microglobulin-MHC antigen dimers as anchorage sites was tested on Daudi human Burkitt lymphoma cells [46, XY, 15q-, 14q+, beta2-m(-), HLA(-)]. The H-Y antigen level of Daudi was only 20% of that of Raji and Ramos, two human male pseudodiploid Burkitt lymphoma lines that were beta2-m(+), HLA(+). When Daudi is hybridized with beta2-m(+), HLA(+) cell lines, beta2-microglobulin, supplied by the latter, is known to restore the expression of Daudi HLA antigens A10 and BW17. Such restoration of HLA antigen expression markedly elevated H-Y antigen levels in those somatic hybrids. Thus the H-Y antigen level of the Daudi x Raji 8A (male X male) hybrid became equal to that of TetraRaji--the colcemide-induced Raji tetraploid line. Two independently derived Daudi x Hela D98 (male x female) hybrids, DAD 1 and DAD 10, demonstrated even higher H-Y antigen levels comparable to that of normal male peripheral blood lymphocytes.


Conservation of the primary structure at the 3' end of 18S rRNA from eucaryotic cells. DNA sequencing methods have been used to determine a sequence of about 20 nucleotides at the 3' termini of various 18S (small ribosomal subunit) RNA molecules. Polyadenylated rRNA was first synthesized using the enzyme ATP:polynucleotidyl transferase from mainze. Then in the presence of an oligonucleotide primer uniquely complementary to the end of each adenylated rRNA, a cDNA copy was produced using AMV reverse transcriptase. In every case, the cDNA transcript was of finite size, which we ascribe to the appearance of an oligonucleotide containing m62A near the 3' end of the 18S rRNAs. Sequences at the 3' termini of 18S rRNA molecules from the four eucaryotic species examined here (mouse, silk worm, wheat embryo and slime mold) are highly conserved. They also exhibit strong homology to the 3' end of E. coli 16S rRNA. Two important differences, however, are apparent. First, the 16S sequence CCUCC, implicated in mRNA binding by E. coli ribosomes, is absent from each eucaryotic rRNA sequence. Second, a purine-rich region which exhibits extensive complementarity to the 5' noncoding regions of many eucaryotic mRNAs appears consistently.


[Isolation and partial characterization of receptor sites of normal human lymphocytes for lectins]. Glycoproteins of the lymphocyte surface are involved in many membrane mediated events. Their specific carbohydrate determinants might interact with lectins. Purification of macromolecules released from normal human lymphocytes by trypsin was performed using gel filtration and affinity properties for Ricinus sanguineus agglutinin. Some structural and biochemical characteristics are given. Relation to HLA determinants and surface immunoglobulins is discussed.


Simultaneous determination of phosphatidylglycerol and the lecithin/sphingomyelin ratio in amniotic fluid. We describe a single-dimension thin-layer chromatographic method by which one can simultaneously determine the lecithin/sphingomyelin ratio and the proportion of phosphatidylglycerol in amniotic fluid. The phospholipids are conveniently detected by an improved molybdenum-blue spray reagent, which immediately produces blue spots on a white background. The stained phospholipids are stable for at least 90 min after spraying, and densitometry results in symmetrical peaks that are easily quantitated.


A rapid radioimmunoassay for human alpha-fetoprotein. Our previous double antibody radioimmunoassay for human AFP was run at 4 degrees C and required 48 h for completion. A method is described which permits the assay to be performed within 5 h due to an increased incubation temperature and the use of polyethylene glycol to precipitate the antigen-antibody complex. The modified radioimmunoassay was standardized against an international reference preparation of human AFP. Characteristics of this assay are discussed.


Placental-like alkaline phosphatase in malignant and benign ovarian tumors. Alkaline phosphatase (ALP) components in extracts of seven malignant and eleven benign ovarian tumors were characterized using the criteria of electrophoretic mobility before and after neuraminidase treatment, heat stability, L-phenylalanine inhibition and reactivity against antiplacental ALP antiserum. Seven of the eighteen tumors had ALP components which most closely resembled the ALP isoenzyme normally found in placenta and were clearly distinguished from all other tissue ALPs. The proportion of tumors with the placental-like ALP in the malignant group (five out of seven) was significantly greater than the proportion in the benign group (two out of eleven). The fraction (78%) of the malignant tumors with the isozyme represents a larger percentage than has previously been found by examination of cancer patients' sera. The electrophoretic mobilities of the placental-like ALPs in the tumors were in no case identical to the mobilities of any of the six common placental ALP phenotypes. The tumor ALPs may thus be determined by rare variant alleles at the ALP locus, or alternatively, the enzyme molecules may have been subject to structural modification. At least two of these tumors contained an electrophoretically slow. heat-stable, leucine-sensitive ALP, which may correspond to what has been termed the D-variant of placental ALP found in some other tumors.


Problems encountered in the measurement of pregnancy-specific beta1-glycoprotein. Evidence is presented here that variants of the human trophoblastic plasma protein, pregnancy-specific beta1-glycoprotein (PSbetaG) are detectable in plasma samples from a small percentage of subjects and may be recognised by their capacity to form indistinct immunoprecipitates and give artificially low levels when measured by radioimmunoassay. Urinary PSbetaG shows a marked degree of heterogeneity and is difficult to measure using conventional immunoprecipitation assay techniques. By radioimmunoassay, late pregnancy urinary PSbetaG concentrations have been found to be in the range 0.35 to 6.9 microgram/ml, approximately 3% of the term plasma concentrations.


Haptenic antibody induced by N-phthaloylated ox insulin in the Hartley guinea-pig. Female Hartley guinea-pigs were immunized with N-phthaloylated ox insulin (96% NA1,NB1,NB29-triphthaloyl ox insulin) using H. pertussis as adjuvant. Haptenic antibody, which was found in antisera taken 20 days after secondary immunization, was assessed for cross-reaction with a series of N-acylated ox insulins. Cross-reaction readily occurred between haptenic antibody and the partial hapten-bearing antigens, NA1-monophthaloyl and NB1-monophthaloyl ox insulins. The immunochemical data suggested that the NA1-glycyl hapten and the NB1-phenylalanyl hapten were both acting as immune determinants for B-lymphocytes, and the determinant competition was taking place between contralateral facets of the principal immunogen, NA1,NB1,NB29-triphthaloyl ox insulin. The data was also consistent with the view that the overall tertiary structure of N-acylated insulins may be very similar to that of their native insulin, the addition of hapten groups having produced only localized areas of structural perturbation.


Drug-induced lupus erythematosus with in vivo lupus erythematosus cells in pleural fluid. Pleural involvement in drug-induced lupus erythematosus is not uncommon. Lupus erythematosus cells were found in vivo in the pleural of an elderly patient who had received procainamide (Pronestyl) hydrochloride (2 gm daily) for nine months. Patients who initially have pleural effusions while receiving drugs capable of inducing lupus erythematosus should have the fluid analyzed for lupus erythematosus cells to help clarify the cause of the effusion.


Factors involved in differential Giemsa-staining of sister chromatids. Microspectrophotometric evaluation of differentially stained sister chromatids made it possible to analyse precisely the factors involved in the Giemsa methods. The concentration of Hoechst 33258, pH of the mounting medium temperature during UV-exposure and the quality (wavelength)of UV-light influenced the differential staining. Exposure of blacklight of 10(-5) M Hoechst 33528-stained brdU-labeled chromosome specimens mounted in McIlvaine buffer (pH 8.0) at 50 degrees C reproducibly allowed differential staining of sister chromatids within 15 min. On the other hand, Korenberg-Freedlender's method using no Hoechst 33258 was also UV-light-dependent. Thus, photolysis of BrdU-substituted DNA was considered the basic mechanism of the Giemsa methods where the photosensitive Hoechst 33258 played a role as a sensitizer.


Proliferation of hepatocytes.

Proliferation of hepatocytes. Hepatocyte proliferation may be controlled by reversible patterns of endocrine changes, monitored by the liver, involving known hormones and their receptors. A two-programme model of related interactions among nutrients, specific lipoproteins, and highly phosphorylated nucleotides is postulated. This hypothesis stems from in vitro studies of rat hepatocyte proliferation under chemically defined conditions and from in vivo studies using partially hepatectomized, hormone-infused, developing and lipotrope-deficient rats. Certain findings are discussed with regard to receptor systems which show negatively cooperative properties; to problems of proliferative specificity; and to novel approaches for defined studies of chemical hepatocarcinogenesis.


[Immune response to sheep erythrocytes fixed in formaldehyde]. Treatment of sheep erythrocytes with 1.5% of formaldehyde alter their immunogenic properties. On the background of immune response to nonfixed erythrocytes, the performed studies showed the following peculiarities in the reaction of mice to fixed erythrocytes: 1) the primary response of animals, in ected with fixed erythrocytes, was rather weak after its determination by the number of rosette forming cells (RFC), antibody forming cells (AFC) in the spleen and by the of hemagglutinins (HA) in serum; 2) in contrast to the primary response the fixed erythrocytes induced a good secondary response, which was considerably higher/according to the number of RFC and titre of HA/in mice sensibilized by nonfixed erytrocytes in advance than in those presensibilized by fixed cells. Therefore the fixed erythrocytes were weaker inducers of immunologic memory than the nonfixed under equal other conditions. 3) In contrast to the nonfixed the fixed erythrocytes did not induce the formation of large amounts of direct AFC during the secondary response as well even then when the animals were presensibilized by nonfixed form of the antigen.


The calculated radial decline of the extracellular action potential compared with in situ measurements in the human brachial biceps. Action potentials from 16 single muscle fibres were recorded with a multi-electrode from the human brachial biceps. Propagation velocity was calculated for 10 of the fibres, and the volume conduction at distances of 79, 152 and 251 micrometer, was measured for 6 of these fibres. The radial decline of the action potentials was calculated from the Fourier transform of the action potential at the origin, simulating the low-pass filter characteristics of the muscle tissue by sections of RC elements. A good agreement was found between the calculated data and the experimental data on the radial decline of the action potentials. The 90% attenuation radius of the action potential recorded with a 25 micrometer electrode, calculated for 16 action potentials, was derived from the volume conduction calculations. The average value found was 191 +/- 20 micrometer. A linear correlation was found between this radius and the peak spectral amplitude of the action potentials. Likewise, a linear correlation between the 90% attenuation radius and the propagation velocity of the fibres was confirmed.


Stretch and Hoffmann reflexes during phasic voluntary contractions of the human soleus muscle. Measurements were made of EMG responses in human soleus muscle, following torque disturbances of the foot during phasic voluntary plantarflexions. These were compared with measurements of the Hoffmann reflex during identical voluntary movements. We observe that stretch adequate to evoke a stretch reflex in the relaxed limb often fails to evoke any response during a phasic contraction until about 120 msec after the onset of the stimulus. During this period the H-reflex is facilitated. These findings are discussed in terms of the servo-theory of stretch reflex function and the concept of trans-cortical stretch reflexes.


Restraint in primate chair may cause unusual behaviour in baboons; electrocorticographic correlates and corrective effects of diazepam. Whereas primates restrained in a chair may usually be considered 'normal', the behaviour of baboons in this position was considerably altered: their eyes were closed and they only slightly reacted to the approach of the experimenter (opening of the eyes), in contrast with their active behaviour when left free to move about in their cage. When the ECoG was recorded from certain cortical areas (particularly somatic area I), an unusual pattern developed during the restrained state: 'drowsiness rhythms', which typically appear in this area during transition from wakefulness to sleep, developed in long sustained sequences. Administration of diazepam, an anxiolytic drug, caused both the behaviour and the ECoG of the restrained animal to return to normal (i.e., attentive behaviour and ECoG characteristic of the true waking state). The prolonged drowsy-like ECoG and behaviour may therefore underline a reaction to the 'stress' conditions brought on by restraint.


Effects of ethanol, caffeine, and placebo on the auditory evoked response. A previous paper (Wolpaw and Penry 1975) described separation of the 75-250 msec portion of the AER into N1P2, a product of large areas of cortex, and the T complex, probably a product of secondary auditory cortex. With monaural stimulation, the T complex is larger and earlier on the side contralateral to stimulation and on the right side. Thirty-one normal adults received 3 oz. of ethanol, 300 mg of caffeine, or placebo. Monaural AERs were recorded before intake in all cases, 1 and 4 h after ethanol and 80 min after caffeine or placebo. Blood levels of ethanol and caffeine were measured. Placebo produced mild (20%) decreases in N1P2 amplitude. Caffeine did not decrease N1P2 amplitude. It did produce a statistically significant 2% decrease in Ta peak latency. Ethanol reduced N1P2 amplitude markedly at 1 h and mildly at 4 h. Placebo did not affect hemispheric differences. Caffeine significantly increased the Ta peak ipsilateral vs. contralateral latency difference in 3 of 7 individuals. Ethanol significantly increased it in 3 of 6 subjects at 1 h and in 7 of 10 at 4 h, primarily by increasing ipsilateral latencies.


Computer-assisted interpretation of clinical EEGs. A multivariate pattern recognition technique has been developed, to distinguish the EEGs of patients with cerebral pathology from those of normal controls and to localize any abnormalities detected. Two methods of feature extraction have been used, power spectral density and slope descriptor analysis, together with various types of feature compression. These techniques have been evaluated on EEGs from 63 patients with proven pathology. Spectral analysis proved more reliable than slope descriptor analysis and predicted the site of cerebral pathology more accurately than did visual assessment of the EEGs. This apparent improvement over the diagnostic reliability of visual analysis in considered to justify further development and evaluation of this technique.


Visually evoked cortical potentials in renal failure: transient potentials. Transient visually evoked cortical potentials (VECPs) were recorded from patients with renal disease. Changes in VECP latencies are described for undialyzed patients, patients receiving dialysis therapy, and patients who received kidney transplants. Characteristics of VECP latencies in these patient groups as well as examples of changes in latency values with respect to time for two individual patients are examined. The basic overall finding is that the VECP latencies increase as a patient's clinical condition deteriorates and normalize as the condition improves.


Late components of average evoked potentials in children with different abilities to concentrate. Three matched groups, each of seven 11-year-old children with good, average and poor ability to concentrate were selected. Evoked potentials and EEG alpha amplitude changes after visual imperative stimuli were studied in warned reaction time experiments. The children with poor ability to concentrate (compared with the other groups) showed longer latencies of the P2 component and reduced amplitudes of the P2-N2 deflection at the vertex. Similar, although attenuated, effects were found in the Fz derivation. No group effects could be established for Oz. These findings are discussed in relation to group differences in pre-stimulus negativity and alpha attenuation, as reported in a previous paper.


Provoked visual impairment in multiple sclerosis studied by visual evoked responses. Pattern-reversal VERs were studied during the visual impairment provoked by exercise in 2 patients with demyelinating optic neuritis. It was found that the transient reduction in visual acuity was correlated  to a transient decrease in the amplitude of the major positive component of the VER, whereas no significant changes could be observed in the latency of the response. The normal VER was not influenced by exercise.


The EEG's of chronic schizophrenic patients in hospital and in the community. EEG's were recorded from chronic schizophrenic inpatients, remitted patients living in the community and normal controls, during resting and auditory stimulation conditions. Chronic impatients had more energy in the slower and fastest wave bands than either remitted patients or normal controls. No differences were found in overall EEG energy and variability between groups of subjects or between sides of the head. It was concluded that any abnormalities were associated with behavioural differences and not with schizophrenic process changes.


Wrist actigraphic measures of sleep and rhythms. With a piezo-electric activity transducer, wrist activity recordings were made simultaneously with EEG, EOG and EMG recordings. Wrist activity alone was used to estimate Total Sleep Time and blind independent scoring of sleep and wakefulness by EEG-EOG-EMG was done. Correlations between the two methods were termined for minutes of Sleep (r = 0.98), Total Sleep Period (r = 0.95) and minutes of Wake Time Within Sleep (r = 0.85). Continuous wrist activity recordings may thus provide inexpensive but very accurate estimates of Sleep Time.


A phase locked loop device for automatic detection of sleep spindles and stage 2. A method is described using phase-locked loops which, as electronic frequency detection devices, provide the basis of a relatively inexpensive system of automatic sleep spindle detection. It is acceptably reliable at quite high rates of data processing.


Synopsis of endoscopic and related morphological findings in Whipple's disease. Whipple's disease is characterized by typical endoscopic lesions in the duodenal wall. They consist of a thickened and inflamed mucosa with yellowish-white spots, in part solitary, in part confluent. Petechial hemorrhages mark the high mucosal vulnerability. Clumsy and dilated villi with ectatic lymph vessels and PAS-positive macrophages are the corresponding histologic feature. In the ultrastructure numerous rod-shaped micro-organisms are evident. Following treatment with tetracycline the mucosal surface normalizes rapidly, the bacteria disappear, whereas other histologic changes remain over a longer period of time.


Gamma-globulins patterns in CSF of inflammatory neurological diseases in tropical Africa. 400 CSF electrophoreses were performed on Black West Africans (Ivory Coast) with inflammatory infections of the nervous system. In the normal state, the CSF of the Black African contains a higher concentration of gamma-globulins than the European (15.8% compared to 8--10%). This study permits us to confirm indirectly the classical idea of the extreme rarity of MS in tropical areas: an increase in the gamma-globulins of the CSF has been shown in 22% of the cases and the two principal responsible infections are trypanosomiasis (57.3%) and SSPE (16.8%). The oligoclonal distribution by various authors in trypanosomiases was not found. On the contrary, a nonclonal aspect of gamma-zone was constantly found (49/50). It is possible that the differences are in accordance with the different evolutionary phases of the disease, and that the oligoclonal distribution is a marker of the autoimmune state of the disease.


Matching for DLA-A, DLA-B and DLA-D antigens and skin allograft survival in unrelated beagle dogs. The effect of matching for immunogenetic markers on the survival of skin allografts in unrelated dogs has been studied. Skin grafts in recipients differing from donors for either DLA-A or DLA-B antigens, or both, had mean survival times (MST) of 9.3, 9.7 and 8.5 days, respectively. Differences between these groups were not significant. Prolonged survival of skin grafts, however, was found in SK-LD-indentical (MST: 12.4 days) or SD-identical/LD-different (MST: 12.5 days) donor-recipient combinations. We conclude that dkin allograft survival in dogs appears to be controlled by DLA-A and DLA-B, but not by DLA-D determinants.


The association between vesical carcinoma and urinary obstruction. Signs of bladder-outflow obstruction on intravenous urography were recorded in 199 male patients with carcinoma of the bladder and in an age-matched control group. The indicence of obstruction was significantly higher in the tumour group (p less than 0.05). Possible explanations for this association are discussed.


On the presence of cysteinesulfinic acid and 2-aminoethanesulfinic acid in human adenomatous prostate. The presence of cysteinesulfinic acid and 2-aminoethanesulfinic acid has never been described in human adenomatous prostate. Results here reported are compared with those in the normal prostate of guinea pigs and rats.


Improved suprapubic vesical drainage in transvesical prostatectomy. An improved drainage is used in transvesical prostatectomy in order to avoid urinary retention after removal of the Novak-type suprapubic drainage with bladder neck cerclage haemostasis (described by some authors in 1975). The causes of urinary retention are analysed, the drainage technique as well as the mechanisms eliminating the above-mentioned complications are described.


The number of antibody-producing clones as revealed by the distribution of allelic variants of immunoglobulin light chains. A new method of evaluation of the number of clonal precursors of antibody-producing cells based on the phenomenon of allelic exclusion is proposed. The distribution of the allotypes of specific antibodies in the population of heterozygous F1 hybrids depends on the number of clonal precursor cells. The smaller the number of precursors, the larger the variance of the distribution. The frequency of clones producing antibodies to the azobenzoate hapten (approximately 10(-8) after first and approximately 10(-9) after second and third immunizations) in (MSU X August)F1 rats calculated in the present work is in reasonable agreement with the data obtained (for other haptens) by limiting dilution cloning assays. The difference in the numbers of anti-hapten and anti-carrier clones is discussed. The numbers of IgM and IgG-synthesizing clones are calculated from the distributions of the allelic variants of corresponding rat antibodies to lactoside. The data suggest the existence of distinct IgG precursors which contradicts the hypothesis of IgM-IgG switch within the clones only.


Human Ia-like DRw lymphocyte antigens stimulating activity in primary mixed lymphocyte reaction. The intensity of the primary mixed lymphocyte reaction (MLR) seems to be influenced by at least two distinct determinants of the HLA-D region: the HLA-Dw stimulating products and the human Ia-like B lymphocyte DR (D-related) antigens. In three families, the primary MLR is significantly higher in case of HLA haploidentity, when stimulating and responding cells differ with regard to both Dw and DRw determinants, than when only Dw products are different. Thus, an additional effect of DR antigens during primary MLR is observed. The role of DRw antigens in primary MLR and the results obtained by a primed lymphocyte test which can discriminate between Dw3 and DRw3, indicate that DRw and Dw products could be distinct determinants.


Regulation of the immune response. I. Regulatory cell equilibrium in the "virgin" state. Spleen cells from pairs of inbred mice (age- and sex-matched) were challenged in vitro with a T-independent antigen. The specific plaque-forming cell (PFC) response in cultures containing cells from both donors was not intermediate to the responses of independently cultured donor spleen cells. Instead, a pattern of significant suppression (67% of pairs tested) or enhancement (22% of pairs tested) was observed. The suppression or enhancement was antigen-specific and did not represent a general phenomenon within the mixed cultures. A suppressive outcome in mixed cell cultures seems to be associated with cells from the donor with the higher individual PFC response. The converse is true for enhancement. The nonadditive nature of the PFC response in mixed cultures and the frequency of suppression to enhancement (3:1) imply that (a) specific, ongoing immunoregulation occurs even in the naive individual, and (b) this regulation is characterized by regular cycles of specific suppression and enhancement.


Protection against fulminant sepsis in splenectomized mice by implantation of autochthonous splenic tissue. Infection of splenectomized mice with Diplococcus pneumoniae, type III, resulted in a fourfold higher mortality rate than did infection of normal mice. Splenectomized animals were protected against fulminant, fatal sepsis by subcutaneous transplantation of autochthonous splenic tissue at the time of splenectomy. Animals with ectopic splenic tissue, and sham-splenectomized control mice, exhibited normal serum opsonin and leukophilic gamma-globulin activity, with respect to pneumococcus, that was lacking in splenectomized animals.


Evidence for the clonal nature of erythropoietic bursts: application of an in situ method for demonstrating centromeric heterochromatin in plasma cultures. Erythropoietic bursts were produced in plasma cultures seeded with a mixture of male and female murine bone marrow cells. Chromosome spreads of C-metaphases were prepared and stained in situ with a modification of the C-banding technique. In cultures seeded with mixtures of male and female cells, homogeneity of male or female C-metaphases in erythropoietic bursts was established by the presence or absence of the Y-chromosome. These results provide evidence that each erythropoietic burst is a clone, and that the erythropoietic burst-forming unit (BFU-E) is a single cell.


The pontocerebellar projection of the uvula in the cat. The occurrence of retrogradely labeled cells in the pontine nuclei was mapped following injections of 0.3-0.4 microliter of a horseradish peroxidase suspension (50% weight/volume) into the uvula (lob. IX of Larsell) in the cat. The uvula was found to receive afferents from three pontine cell collections. One of these is situated in the paramedian pontine nucleus close to the midline. It forms a fairly distinctly outlined longitudinal column of cells and is present at all levels of the pons except most rostrally and caudally. Another group, in the dorsolateral and lateral pontine nuclei, extends as a somewhat shorter cell column in the longitudinal direction. The third region consists of cells within the rostral part of the peduncular nucleus in its dorsomedial region. The pontine projection to the uvula is bilateral, with some preponderance of crossed connections. The projection to the uvula is organized according to the pattern determined previously for pontine projections to other parts of the cerebellum. A single lobule or part of its receives afferents from more than one cell group in the pons. The projecting cells are most often arranged in longitudinal columns. Correlations with data on the termination of afferents to the pons permit some conclusions regarding the sources of information reaching the uvula via the pons. Main sources seem to be the superior and inferior colliculi, the intracerebellar nuclei and the sensorimotor cortices.


Subcortical afferents to the prefrontal cortex in rabbits. The origins of cells projecting to the prefrontal cortex of the rabbit were studied, using horseradish peroxidase (HRP) technique. HRP injected into the prefrontal cortex labeled cells in the basal forebrain, lateral hypothalamus, raphe nuclei and locus coeruleus area on both sides. Labeled cells appeared also in the nucleus medialis dorsalis of the thalamus and other thalamic nuclei on the injection side.


Identification of lymphocyte subpopulations by simultaneous E-rosette formation and unspecific acid esterase staining. E-rosettes of human peripheral blood lymphocytes were stained by an unspecific acid esterase stain; 74.5% of the rosette forming cells were esterase positive, while only 30% of the non-rosette forming cells showed a reaction product.