Biomedical subjects

R L Allen

Publications and source records attributed to R L Allen.

At least 55 records · Page 3Linked to original sources

PubMed1988

A novel hsp70-like protein (P70) is present in mouse spermatogenic cells.

Mouse spermatogenic cells contain relatively large amounts of a 70-kilodalton protein (P70) that is closely related to hsp70, the major inducible heat shock protein. When hsp70 from spermatogenic cells is heat induced, it migrates to the same location as does P70 on two-dimensional polyacrylamide gels, indicating that it has an apparently identical mass and isoelectric point. P70 reacts strongly and specifically with an anti-Drosophila hsp70 monoclonal antibody that is specific for products of the hsp70 gene family. Both P70 and hsp70 are also ATP-binding proteins and are purified by using ATP-affinity chromatography. However, P70 and hsp70 are unique proteins on the basis of peptide map analysis and are regulated differently in germ cells. P70 appears to be a novel heat shock protein of spermatogenic cells which is synthesized in association with germ cell differentiation.

Animals↗

PubMed1988

Expression of heat shock proteins by isolated mouse spermatogenic cells.

Proteins of the hsp70 family are abundant in mouse spermatogenic cells. These cells also synthesize relatively large amounts of a 70,000-molecular-weight protein (P70) that appears to be a cell-specific isoform of hsp70, the major heat-inducible protein (R.L. Allen, D.A. O'Brien, and E.M. Eddy, Mol. Cell. Biol. 8:828-832, 1988). In this study, proteins of unstressed and heat-stressed spermatogenic cells consisting of purified preparations of preleptotene, leptotene-zygotene, pachytene spermatocytes, and round spermatids were analyzed by two-dimensional polyacrylamide gel electrophoresis. Unstressed preleptotene and leptotene-zygotene spermatocytes contained little P70, whereas relatively large amounts of P70 were present in pachytene spermatocytes and round spermatids. Labeling studies showed that P70 was synthesized primarily in pachytene spermatocytes and that little synthesis occurred in round spermatids or in preleptotene and leptotene-zygotene stages of spermatogenesis. Synthesis of hsp70 was not detectable in unstressed cells but was induced in all stages of isolated germ cells following heat stress. These results indicate that P70 is expressed in a stage-specific manner during cell differentiation, whereas hsp70 is synthesized in response to stress in all populations of isolated spermatogenic cells examined.

Animals↗

PubMed1987

Molecular cloning, characterization, and complete nucleotide sequence of the gene for pneumolysin, the sulfhydryl-activated toxin of Streptococcus pneumoniae.

A recombinant lambda bacteriophage has been isolated that carries DNA from Streptococcus pneumoniae and expresses a potent hemolysin that has been shown to be pneumolysin, the sulfhydryl-activated toxin of the pneumococcus. Hemolytic activity is inhibited by cholesterol and neutralized by serum against streptolysin O. The cloned gene expresses two polypeptides (Mrs, 56,000 and 53,000) in an Escherichia coli in vitro transcription-translation system, and both are precipitated by the addition of anti-alveolysin serum and anti-streptolysin O serum in the presence of Staphylococcus aureus cells. Expression of pneumolysin occurs when the gene is cloned in both possible orientations in pUC8. The DNA sequence of a 5-kilobase ClaI fragment that carries the pneumolysin gene has been determined. An open reading frame was identified that encodes a polypeptide of 471 amino acids that is hydrophobic in character and has an N-terminal amino acid sequence which is identical to that deduced from amino acid sequencing of the purified protein. The predicted amino acid sequence of the polypeptide reveals a single cysteine residue located 44 residues from the C terminus. Putative promoter and ribosome binding sites have been identified 5' to the pneumolysin coding sequence.

Amino Acid Sequence↗

PubMed1986

Phaeohyphomycosis complicating compound skull fracture.

Intracranial infection is a well-recognized complication of compound skull fractures. In most cases various bacteria are identified as the etiologic agents. Fungal infection complicating open head trauma is unusual. We describe a patient who contracted fatal meningoencephalitis due to Drechslera spicifera, a rarely pathogenic soil saprophyte, after open head trauma.

Adult↗

PubMed1986

A comparative study of hepatic and epidermal histidase in the guinea-pig (Cavia porcellus).

Histidine ammonia lyase was purified to homogeneity from guinea-pig liver and epidermis. Both enzymes had similar molecular weights, subunit composition and pH optima. Km values for the two were similar at pH 9.2 but different at pH 7.0. Both enzymes were stimulated by low thiol concentrations and inhibited at higher concentrations, but to different extents. Antibody to the hepatic enzyme showed complete identity against hepatic enzyme but incomplete identity against epidermal enzyme.

Ammonia-Lyases↗

PubMed1986

Examination of the macronuclear replication band in Euplotes eurystomus with monoclonal antibodies.

A panel of eight monoclonal antibodies (MAbs) was prepared from spleen cells of mice immunized with macronuclear replication bands (RBs) isolated from Euplotes eurystomus. Antibodies were investigated with a solid phase radioimmunoassay (RIA) using either soluble chromatin from isolated RBs or from total macronuclei as antigen. The RIA showed that several MAbs recognized antigens present only in the RB or macronucleus, whereas others recognized antigens present in both structures. Specificity of the MAbs was also examined by indirect immunofluorescence. Antibody C10 recognized an antigen in the rear zone of the RB, whereas MAbs G6 and B2 appeared to stain both the forward and rear zones of the RB. Antibody A7 recognized an epitope distributed throughout the macronucleus except in the RB. Cytochemical studies with degradative enzymes suggested that antigens localized by immunofluorescence were composed of proteins. Immunoblots of SDS PAGE permitted identification of a few proteins that reacted with three of the RB-specific MAbs. Monoclonal antibodies that identify the presence or absence of reactivity of specific proteins in the RB could prove useful in the study of chromatin structure and the mechanism of chromatin replication.

Animals↗

PubMed1986

Brain peptides and glial growth. I. Glia-promoting factors as regulators of gliogenesis in the developing and injured central nervous system.

Glia-promoting factors (GPFs) are peptides of the central nervous system which accelerate the growth of specific glial populations in vitro. Although these factors were first discovered in the goldfish visual system (Giulian, D., Y. Tomozawa, H. Hindman, and R. Allen, 1985, Proc. Natl. Acad. Sci. USA., 83:4287-4290), we now report similar peptides are found in mammalian brain. The cerebral cortex of rat contains oligodendroglia-stimulating peptides, GPF1 (15 kD) and GPF3 (6 kD), as well as astroglia-stimulating peptides, GPF2 (9 kD) and GPF4 (3 kD). The concentrations of specific GPFs increase in brain during periods of gliogenesis. For example, GPF1 and GPF3 are found in postnatal rat brain during a peak of oligondendroglial growth while GPF2 and GPF4 are first detected at a time of astroglial proliferation in the embryo. Stab wound injury to the cerebral cortices of rats stimulates astroglial proliferation and induces marked elevations in levels of GPF2 and GPF4. Our findings suggest that two distinct classes of GPFs, those acting upon oligodendroglia and those acting upon astroglia, help to regulate cell growth in the developing and injured central nervous system.

Animals↗

PubMed1985

Observations on the in vitro formation, development and differentiation of trilaminar vesicles formed from enzymatically dispersed porcine blastocysts.

The formation of trophoblastic vesicles and their use as a model for cell-to-cell interactions in the embryonic development of the mouse have been extensively studied both in vitro and in vivo . In this report, we describe the formation of trilaminar vesicles in vitro from enzymatically dispersed porcine blastocysts. Trilaminar vesicles were generated from single cell suspensions of enzymatically dispersed porcine blastocysts recovered 12 through 15 days post-breeding. Blastocysts at 10 and 11 days of development failed to form TV. No preference for self-aggregation was apparent as evident by the formation of vesicles containing cells from two different blastocysts origins. Two distinct morphological types of vesicles were formed: vesicles that displayed complete homogeneity in cell density and vesicles that had a small area of greater cell density resembling the embryonic disc. Some TV displayed localized areas of nucleated eosinophilic cells within the mesenchymal layer of the trilaminar membrane. In rare instances (2%) TV displayed contractility associated with a raised area of cells on the outer surface of the membrane. The cells comprising this contractile region were positive for Ca(++)-activated myofibriliary ATPase acitivity. Transfer of TV to the uterine horn of a gilt failed to delay a return to estrus. Transfer of TV to the uterine horn of pseudopregnant rabbits induced a localized inflammation and infiltration of leukocytes three days after transfer but no inflammation was evident at 9 or 16 days post-transfer.

Journal Article↗

PubMed1985

Peptides from regenerating central nervous system promote specific populations of macroglia.

The regenerating central nervous system of goldfish contains peptides referred to as glia-promoting factors (GPFs) that stimulate the proliferation of mammalian macroglia. We find that, in vitro, GPF1 and GPF3 promote the appearance of oligodendroglia and GPF2 and GPF4 stimulate proliferation of astroglia. The activities of GPF1, GPF3, and GPF4 increase during regeneration of the goldfish visual system. These results suggest that brain peptides may play a role in the recovery of the injured central nervous system by regulating the growth and development of specific macroglial populations.

Animals↗

PubMed1985

Evaluation of acute nonpenetrating cervical spinal cord injuries with CT metrizamide myelography.

Computerized tomography metrizamide myelography was performed in 46 patients with acute, nonpenetrating cervical spinal cord injuries. By visualizing the spinal canal, spinal cord, and any compressive lesion, the study proved valuable in the decision as to whether surgical decompression was indicated and what approach should be used. Eleven patients were found to have significant spinal cord compression, 10 of whom were treated surgically. The technique, results, and complications resulting from the study are discussed.

Adolescent↗

PubMed1985

Isolation and characterization of chromatin replication bands and macronuclei from Euplotes eurystomus.

A method is described for isolating replication bands (RBs) from Euplotes eurystomus in quantities sufficient for biochemical analysis. The method involves the disruption of whole cells in a low ionic strength buffer that maintains RB integrity while dispersing macronuclear chromatin. The RBs are then stabilized with MgCl2 and spermidine phosphate and isolated by gradient centrifugation. Staining with silver nitrate and thiol-specific coumarin maleimide has been demonstrated in the RBs of Euplotes and other hypotrichs; both of these properties were maintained in isolated RBs. A method is also described in this study for isolating highly purified macronuclei. Examination of isolated macronuclei and RBs with electron microscopy (EM) indicates that the morphology of both structures remain essentially intact during purification. We also observe with EM an increase in the number of replicating molecules in RBs compared to macronuclei. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) demonstrates a consistent but minor enrichment of a 55 kilodalton protein in RBs when compared to macronuclear proteins.

Animals↗

PubMed1985

Distribution of intrarenal blood flow consequent to left atrial balloon inflation.

The effects of inflation of a balloon within, and consequent distension of, the left atrium (LABI, left atrial balloon inflation) on total renal blood flow (RBF) and intrarenal blood flow distribution were measured and compared to values obtained from another group of dogs that were hemorrhaged (HEM) to the same level of hypotension as that produced by LABI, a mean systemic arterial pressure of 88 mm Hg. Kidney wt/kg, RBF/kg body wt, and urine flow were markedly reduced during the hemorrhage period in the HEM group when compared to values obtained during the experimental period for the LABI group. Data from the freeze-dissection (133Xe) analysis revealed that the percentage distribution of blood flow as renal outer cortical (OC) blood flow was less (26%) in the HEM group than in the LABI group (50%), this latter value being very similar to that of control dogs that experienced no hypotension (49%). LABI better maintains OC blood flow and urine flow when compared to HEM at the same systemic blood pressure, suggesting a role for cardiopulmonary receptors in reflex sympathetic control of renal blood flow distribution during hypotension.

Animals↗

PubMed1984

Cytochemistry of the chromatin replication band in hypotrichous ciliated protozoa staining with silver and thiol-specific coumarin maleimide.

A modification of the silver-staining techniques for nucleolar organizing regions (NORs) was used to stain selectively the macronuclear replication bands (RBs) and nucleoli in hypotrichous ciliated protozoa (Euplotes, Stylonychia, and Oxytricha). Silver staining of both types of structures was trypsin-sensitive and DNase I-insensitive, suggesting the involvement of proteins. Silver-staining proteins in the RB were differentially extracted with acid, without any decrease in nucleolar staining. Triton-acid-urea gel electrophoresis of an acid extract of Euplotes macronuclei revealed enhanced silver reaction with a single protein upon selective silver staining. An abundance of thiol groups was also demonstrated in the RBs and nucleoli by the fluorochrome 3-(4-maleimidylphenyl)-7-diethylamino-4-methyl coumarin (coumarin maleimide). Histochemical studies, including blocking thiols with N-ethyl maleimide (NEM), indicated that thiols were not necessary for silver staining, and that proteins in the RBs and nucleoli reacting with coumarin maleimide were not acid extractable.

Animals↗

PubMed1984

One-step sucrose dilution of frozen-thawed sheep embryos.

Sheep embryos of the late morula to early blastocyst stage were frozen, thawed and cultured to test several sucrose solutions for post-thaw dilution of the cryoprotective agent glycerol. Ewes of mixed breeding were superovulated and embryos were flushed from the uterus either surgically or at slaughter 5 d after estrus. Fifty-eight embryos were pooled in microdrops of modified Dulbecco's phosphate buffered saline (MDPBS) then randomly divided into four treatments. A 2 x 2 factorial design was used to compare 0.25 M sucrose in MDPBS as an in-straw cryoprotectant dilution with a standard step-wise dilution procedure within standard fast and slow freeze-thaw systems. After storage in liquid nitrogen for 6 to 8 d, the embryos were thawed and the cryoprotectant (1.4 M glycerol) removed before culture in microdrops of modified synthetic oviduct fluid under paraffin oil in water-saturated 5% CO(2) in air atmosphere at 37 C. No significant interaction was found between the freeze-thaw procedure and cryoprotectant + dilution procedures. Embryos in the fast freeze-thaw procedure had a mean development score of 1.3 +/- 0.3 and those in the slow freeze-thaw procedure had a mean score of 1.2 +/- 0.3. The mean development score 2.0 +/- 0.3 for the standard dilution procedure was superior (P<0.001) to the score of 0.6 +/- 0.2 for the 0.25 M sucrose dilution procedure. In a separate trial, 18 sheep morulae were collected and equilibrated with 1.4 M glycerol in MDPBS. A standard fast freeze-thaw procedure was used and, after 18 d of storage at -196 C, the glycerol was diluted from the embryo with 1.0 M sucrose. Culture was conducted in a similar manner and a mean development score of 1.0 +/- 0.3 was obtained. These results indicate standard cryoprotectant dilution procedures for sheep embryos are superior to dilution with 0.25 M sucrose. In a limited study, dilution with 1.0 M sucrose was also not as effective as standard dilution procedures.

Journal Article↗

PubMed1984

Proteolytic processing of h1-like histones in chromatin: a physiologically and developmentally regulated event in Tetrahymena micronuclei.

Micronuclei isolated from growing cells of Tetrahymena thermophila contain three H1-like polypeptides alpha, beta, and gamma. Micronuclei isolated from young conjugating cells (3-7 h) also contain a larger molecular weight polypeptide, X, which is being actively synthesized and deposited into these nuclei (Allis, C. D., and J. C. Wiggins, 1984, Dev. Biol., 101:282-294). Pulse-chase experiments (with growing and conjugating cells) suggested that X is a precursor to alpha and that alpha is further processed to gamma and a previously undescribed and relatively minor species, delta. These precursor-product relationships were supported by cross-reactivity with polyclonal antibodies raised against alpha and peptide mapping. While beta consistently became labeled under chase conditions (both in growing and mating cells), it was not clear whether it is part of the vivo processing event(s) which interrelates X, alpha, gamma, and delta. Beta was not recognized by alpha antibodies. Despite this uncertainty, these results suggest that proteolytic processing serves to generate significant changes in the complement of H1-like histones present in this nucleus.

Animals↗

PubMed1984

In vitro development of porcine embryos in coculture with endometrial cell monolayers or culture supernatants.

The objective of this study was to determine if porcine endometrial cell contact would enhance the in vitro development of porcine embryos relative to endometrial cell culture supernatants or ovarian fibroblast contact. One hundred forty-nine embryos of various developmental stages were collected from 10 excised gilt reproductive tracts. Embryos were washed three times and placed randomly into one of four treatments: Minimal Essential Medium + 5% heat-treated fetal calf serum (complete MEM), porcine endometrial cell coculture (pEM), porcine endometrial cell supernatant (pEMS) and porcine ovarian fibroblast coculture (pOF). All cultures were conducted in 96 well microtiter plates, one embryo per well, at 37 C in a 5% CO2 in humidified air atmosphere. Observations for stage of development were made every 12 h by phase contrast microscopy (100X). Embryos in the pEM and pOF treatments had significantly greater (P less than .05) mean embryonic development scores (1.59 +/- .22 and 1.44 +/- .18, respectively) than embryos cultured in complete MEM only or pEMS (.63 +/- .13 and .48 +/- .15, respectively). The results of this study indicate that cell to embryo contact enhances in vitro development of porcine embryos.

Animals↗

PubMed1984

Adolescents' communication styles and learning about birth control.

Adolescents have different communication styles for acquiring information about birth control from a mass medium--interactant or noninteractant, depending on whether they involve communication with others in the process of media use. Three interactant styles are identified: a home-oriented style, where communication is primarily with immediate members and friends; a peer-oriented style, where communication is primarily with own-age peers and personal friends; and multi-source user, where communication is with more than one cluster of sources and the majority of one's communication is not with any particular source. The noninteractant style identified is a media-oriented style, where the adolescent relies exclusively on the mass media for information acquisition without communicating with anyone about the content or process of learning. Data suggest that the interactant style, especially for the multi-source user, is most beneficial for new learning. Gender and the presence of siblings at home are important moderators of the relationship between communication styles and knowledge of birth control. For example, females with a media-oriented style know more about birth control than females with a home-oriented style. Implications of the findings for the delivery of birth control information to adolescents are discussed.

Adolescent↗

PubMed1983

Sucrose dilution of frozen mouse embryos: interaction of glycerol and sucrose concentrations.

Several concentrations of glycerol for cryoprotection and several concentrations of sucrose for cryoprotectant dilution were examined with frozen, thawed and cultured mouse embryos. Four hundred and eighty late morulae to early blastocyst stage embryos were collected from 35 superovulated mice (B6D2 x Swiss Webster crosses back-crossed to Swiss Webster males) 3-1/2 days after breeding. The embryos were transferred through increasing concentrations of glycerol in modified Dulbecco's phosphate buffered saline (MDPBS) to reach three final concentrations of 1.0 M, 1.4 M and 1.8 M. The embryos were loaded in 0.5-ml French straws appropriately filled with the cryoprotectant and sucrose solutions for each treatment. The straws were cooled with a standard fast-freezing program to -35 degrees C, then plunged into liquid nitrogen. After 58 days of storage at -196 degrees C the straws were thawed in a 37 degrees C water bath. Cryoprotectant dilution was accomplished with a standard step-wise procedure or in the straw with one of three concentrations of sucrose solution (0.25 M, 0.5 M, 1.0 M) in MDPBS. The embryos were then washed twice in MDPBS, twice in Whitten's media for embryo culture and then placed in microdrops of Whitten's media under paraffin oil in a water saturated 5% CO2 in air atmosphere at 37 degrees C. Embryos were observed 24 hours later for development to the expanded blastocyst stage. The proportion of embryos developing in vitro from the three glycerol concentrations were not significantly different with standard step-wise dilution procedures for glycerol removal. After step-wise cryoprotectant removal, blastocyst expansion occurred in 49%, 44% and 52% of embryos frozen in 1.0 M, 1.4 M and 1.8 M glycerol, respectively. The 1.0 M sucrose dilution of 1.0 M glycerol showed the highest development (60.5%) in vitro but was not significantly different from any of these three step-wise diluted glycerol concentrations. The step-wise dilution of the three glycerol concentrations and dilution of the 1.0 M glycerol and 1.0 M sucrose were all superior (P<0.01) to any other dilution procedure examined.

Journal Article↗