The influence of therapy (sympathomimetic amines versus vasodilators) in the indexes of ejection fraction derived from an analysis of Doppler aortic flow

The purpose of this study was to map the distribution of alpha-subunits of G-proteins--Galpha(olf s), Galpha(olf), Galpha(s), Galpha(i), Galpha(o), Galpha(z) and Galpha(q11)--in developing, denervated or injured rat molar teeth, using fluorescence microscopic immunohistochemistry coupled with immunogold electron microscopic immunocytochemistry. In rat fetuses (E17-E21), a widespread expression of Galpha(q11) was seen in maxillary/mandibular mesenchyme as well as in developing teeth. In addition, intensely Galpha(o)-positive nerve fibers were associated with the dental epithelium and the dental papilla of developing teeth. Other G proteins were absent or sparsely distributed during early tooth development. In the adult tooth pulp, odontoblasts appeared to express mainly Galpha(olf s), Galpha(o), and Galpha(q11). Nerve fibers were immunoreactive to Galpha(i), Galpha(o) and Galpha(z). In addition, pulpal blood vessels expressed varying levels of Galpha(olf s) Galpha(z) and Galpha(q11) while Galpha(olf s), Galpha(olf), Galpha(o) and Galpha(q11) were found in various pulpal mesenchymal cells. After adult denervation, nerve fiber-related G-protein immunoreactivity disappeared, but no other changes in pulpal G-protein immunoreactivity were noted. Odontoblasts and mesenchyme cells were intensely Galpha(i)-positive underneath a pulpal traumatic exposure, indicating an injury-induced pulpal upregulation of Galpha(i). The findings that Galpha(i), Galpha(o) and Galpha(z) are expressed in pulpal sensory nerve fibers suggest that these G proteins participate in signal conveyance from the target to the trigeminal nerve cell body.     

Detection of antibodies against equine herpesvirus types 1 and 4 by using recombinant protein derived from an immunodominant region of glycoprotein B

The N-terminal fragment comprising residues +1 to +50 (gB1-50) of equine herpesvirus type 1 (EHV-1) glycoprotein B was expressed as a glutathione S-transferase fusion protein in Escherichia coli. Recombinant gB1-50 (rgB1-50) was recognized in immunoblots by sera from rabbits immunized with EHV-1 and by convalescent-phase sera from horses with natural EHV-1 infections. An enzyme-linked immunosorbent assay (ELISA) for monitoring antibody levels against EHV-1 was developed by using rgB1-50, and its specificity was assessed with a panel of reference antisera against other equine viruses. A specific cross-reaction was detected with EHV-4, which was confirmed by inhibition ELISA. Convalescent-phase sera from horses with natural EHV-1 or EHV-4 infections possessed antibody titers against rgB1-50 ranging from 1:2,000 to 1:64,000, indicating the presence of an immunodominant antigenic site. The study demonstrated the potential application of rgB1-50 as a diagnostic antigen and highlights the glutathione S-transferase fusion system as a simple and effective method of producing purified milligram quantities of antigen.     

Tyrosine phosphorylation of the novel protein-tyrosine kinase RAFTK during an early phase of platelet activation by an integrin glycoprotein IIb-IIIa-independent mechanism

A key regulatory event controlling platelet activation is mediated through the phosphorylation of several cellular proteins by protein-tyrosine kinases. The related adhesion focal tyrosine kinase (RAFTK) is a novel cytoplasmic tyrosine kinase and a member of the focal adhesion kinase (FAK) gene family. FAK phosphorylation in platelets is integrin-dependent, occurs in a late stage of platelet activation, and is dependent on platelet aggregation. In this study, we have investigated the involvement of RAFTK phosphorylation during different stages of platelet activation. Treatment of platelets with thrombin induced, in as early as 10 s, a rapid tyrosine phosphorylation of RAFTK in a time- and concentration-dependent manner. Treatment of platelets with thrombin in the absence of stirring or pretreatment of platelets with RGDS peptide prevented platelet aggregation, but not RAFTK phosphorylation. Furthermore, phosphorylation of RAFTK did not require integrin engagement since platelets treated with the 7E3 inhibitory antibodies that block fibrinogen binding to glycoprotein IIb-IIIa did not inhibit RAFTK phosphorylation. Similarly, platelets treated with LIBS6 antibodies, which specifically activate glycoprotein IIb-IIIa, did not induce RAFTK phosphorylation. Stimulation of platelets by several agonists such as collagen, ADP, epinephrine, and calcium ionophore A23187 induced RAFTK phosphorylation. Tyrosine phosphorylation of RAFTK in platelets is regulated by calcium and is mediated through the protein kinase C pathway. Phosphorylation of RAFTK is dependent upon the formation of actin cytoskeleton as disruption of actin polymerization by cytochalasin D significantly inhibited this phosphorylation. The RAFTK protein appears to be proteolytically cleaved by calpain in an aggregation dependent manner upon thrombin stimulation. These results demonstrate that RAFTK is tyrosine-phosphorylated during an early phase of platelet activation by an integrin- independent mechanism and is not dependent on platelet aggregation, suggesting different mechanisms of regulation for FAK and RAFTK phosphorylation during platelet activation.     

Regulation of the acute phase response genes alpha 1-acid glycoprotein and alpha 1-antitrypsin correlates with sensitivity to thermal injury

BACKGROUND: The response to thermal injury is a complex physiologic process requiring communication between sites of injury and distal target organs. The liver, one of these target organs, synthesizes a family of secretory proteins, the acute phase reactants (APRs), that carries out specific protective functions. This study investigates the response of positively regulated (alpha 1-acid glycoprotein and alpha 1-antitrypsin) and negatively regulated (albumin) APR genes to severe thermal injury in three rat strains with differing abilities to survive thermal stress. METHODS: Age and weight matched male Buffalo, Sprague-Dawley, and Fischer 344, 12- to 16-week-old rats (275 to 325 gm) received a 40% total body surface area scald burn. Total RNA was isolated from livers at 0, 2, 5, 12, 24, and 48 hours. Northern blot hybridization was performed with 32P-labeled rat alpha 1-glycoprotein, rat albumin, and mouse alpha 1-antitrypsin cDNAs. Relative amounts of alpha 1-glycoprotein, alpha 1-antitrypsin, and albumin mRNAs were determined by means of densitometric analyses. RESULTS: All three strains elicit both a positive and negative acute phase (AP) response. Significant differences were observed in the degree and kinetics between strains. Those more sensitive to thermal injury exhibited a more intense positive AP response and possibly a delayed recovery. The AP response between these strains correlates with the variation in ability to survive severe trauma. CONCLUSIONS: The differences in the kinetics and intensity of induction of APR genes between Buffalo, Sprague-Dawley, and Fischer rat strains suggest that the least intense AP response and its timely recovery correlated with the ability to survive a severe thermal injury and that, conversely, the more intense and prolonged response correlated with sensitivity to severe thermal injury. We propose that this may be a basis for variation in survival to thermal injury.     

Structural features of an arabinan fragment isolated from the water-soluble fraction of dehulled rapeseed

A water-soluble polysaccharide fraction was prepared from dehulled rapeseed meal (winter rapeseed variety Casino). Further purification yielded two major fractions having a high content of arabinose and galactose residues, with Ara/Gal ratios of 5.4 (G1) and 1.8 (G2). The Ara/Gal ratio of the high molecular weight fraction G1 was stable over the whole gel filtration peak, indicating that the arabinose and galactose residues are part of the same polysaccharide. The high molecular weight fraction G1 was studied further by methylation analysis and several NMR techniques. Structural studies showed G1 to consist mainly of arabian fragments, which have terminal alpha-L-arabinofuranosyl groups with anomeric carbons bound (1-->5) (A) or (1-->2) (B), and 2,5-substituted arabinosyl residues with anomeric carbons bound (1-->5) (D) or (1-->2) (C) to adjacent arabinosyl residues. The A:B:C:D ratios were 2:1:1:1 according to results from NMR and methylation analysis.     

Inhibitory effect of magnosalin derived from Flos magnoliae on tube formation of rat vascular endothelial cells during the angiogenic process

An aqueous water extract of Flos magnoliae, a Japanese Sino-medicine, inhibits angiogenesis in adjuvant-induced mouse pouch granuloma. Magnosalin (MSA) and magnoshinin (MSI), neolignans isolated from magnolia, have a crucial role in the anti-angiogenic effect of magnolia (Kimura et al., Int. Arch. Allergy Appl. Immunol., 93, 365 (1990); Phytother. Res., 6, 209 (1992)). We investigated the effects of these neolignans on tube formation of endothelial cells (EC) cultured in type I collagen gel during the angiogenic process. MSA (0.1-10 microM), MSI (0.23-7 microM) and corticosterone (CS: 0.3-30 microM) inhibited fetal bovine serum (FBS)-stimulated tube formation in a concentration-dependent manner. Their 30% inhibitory concentration (IC30, 95% confidence limits) values were 0.51 (0.20-1.27) for MSA, 8.14 (2.48-26.7) for MSI and 3.65 microM (2.47-5.40) for CS, respectively. MSA and MSI (1-3 microM) also inhibited interleukin (IL)-1 alpha-stimulated tube formation in a concentration-dependent manner. Their IC50 values (95% confidence limits) were 1.22 (1.01-1.47) for MSA and 0.74 microM (0.24-2.31) for MSI against a submaximal concentration (69 pM) of IL-1 alpha-stimulated tube formation. Their inhibitory effects on the action of IL-1 alpha were non-competitive. These results demonstrate that MSA inhibited FBS-stimulated tube formation with a greater potency than MSI. The inhibitory effect of MSA on the action of FBS differed from that on the action of IL-1 alpha.     

Purification and characterization of diacylglycerol acyltransferase from the lipid body fraction of an oleaginous fungus

BACKGROUND AND PURPOSE: Ischemic cerebral infarction (CI) is a serious complication of acute myocardial infarction (MI). Little information exists on CI after thrombolytic therapy for MI. METHODS: Of 3924 MI patients treated with recombinant tissue plasminogen activator (rt-PA) and heparin, 29 (0.7%) developed CI after treatment. All CI patients had detailed neurological evaluations, and 27 (93%) had CT scans centrally reviewed. RESULTS: Age range was 40 to 74 years (mean, 60 years); 25 patients (86%) were men, and 22 (76%) were white. The electrocardiographic location of MI was anterior in 22 (76%) and nonanterior in 7 (24%). Five CIs occurred within 6 hours, 4 between 6 to 24 hours, 8 during the remainder of the first week, 10 during the second week, and 2 others distributed over the 4 weeks after study entry. Six of 29 CIs did not involve the cerebral cortex; 9 patients (31%) had multiple CIs. Of 28 CIs thought to be embolic in origin, 17 showed strong evidence for at least one cardiac abnormality (mural clot, wall-motion abnormality, aneurysm, or atrial fibrillation) known to be associated more specifically with embolism than MI. Eight of 27 CIs (30%) with CT scans had hemorrhagic transformation of varying degrees; 5 were symptomatic. CONCLUSIONS: The time of occurrence and sites of CI after rt-PA and heparin therapy for acute MI are similar to those reported during the prethrombolytic era.     

Molecular mechanisms for the induction of chromosomal aberrations in CHO cells heated in S phase

During the past 20 years, 972 microvascular transplantations have been performed for 783 patients, with an overall failure rate of 6.2 percent. Fifty-four of the 60 failed transplantations were available for long-term follow-up and were retrospectively reviewed with respect to the original indications for transplantation, the number, and the type of salvage procedures performed following transplant failure. This study illustrates that the choice of salvage procedures performed following transplant failure depends on the original indications, the location, and the severity of the resultant wound. Failure following transplantation for coverage of contour defects or unstable wounds can often be managed by non-microsurgical methods. In contrast, when the indications for transplantation included the transfer of specialized tissues for thumb or digit reconstruction, the restoration of motor or sensory function, or the coverage of a limb-threatening wound, requirements for reconstruction could be satisfied only by a second successful tissue transplant. Eighteen of the 54 cases underwent an additional transplantation, with an 89 percent success rate.     

Targeting of a short peptide derived from the cytoplasmic tail of the G1 membrane glycoprotein of Uukuniemi virus (Bunyaviridae) to the Golgi complex

Members of the Bunyaviridae family acquire an envelope by budding through the lipid bilayer of the Golgi complex. The budding compartment is thought to be determined by the accumulation of the two heterodimeric membrane glycoproteins G1 and G2 in the Golgi. We recently mapped the retention signal for Golgi localization in one Bunyaviridae member (Uukuniemi virus) to the cytoplasmic tail of G1. We now show that a myc-tagged 81-residue G1 tail peptide expressed in BHK21 cells is efficiently targeted to the Golgi complex and retained there during a 3-h chase. Green-fluorescence protein tagged at either end with this peptide or with a C-terminally truncated 60-residue G1 tail peptide was also efficiently targeted to the Golgi. The 81-residue peptide colocalized with mannosidase II (a medial Golgi marker) and partially with p58 (an intermediate compartment marker) and TGN38 (a trans-Golgi marker). In addition, the 81-residue tail peptide induced the formation of brefeldin A-resistant vacuoles that did not costain with markers for other membrane compartments. Removal of the first 10 N-terminal residues had no effect on the Golgi localization but abolished the vacuolar staining. The shortest peptide still able to become targeted to the Golgi encompassed residues 10 to 40. Subcellular fractionation showed that the 81-residue tail peptide was associated with microsomal membranes. Removal of the two palmitylation sites from the tail peptide did not affect Golgi localization and had only a minor effect on the association with microsomal membranes. Taken together, the results provide strong evidence that Golgi retention of the heterodimeric G1-G2 spike protein complex of Uukuniemi virus is mediated by a short region in the cytoplasmic tail of the G1 glycoprotein.     

N-nitroso-N-methylvinylamine: reaction of the epoxide with guanyl and adenyl moieties to yield adducts derived from both parts of the molecule

N-Nitroso-N-methylvinylamine was synthesized and treated with dimethyldioxirane to produce 1-(N-nitrosomethylamino)oxirane. 1-(N-Nitrosomethylamino)oxirane had a t1/2 of < 5 s in buffer at neutral pH and 23 degrees C. This epoxide reacted with Ado to form 1,N6-etheno(epsilon-)Ado. It also reacted with DNA to form products arising from the oxirane portion of the molecule [N7-(2-oxoethyl)Gua,N2,3-epsilon-Gua, and 1,N6-epsilon-dAdo] and the methyl group (N7-methylGua). NADPH-fortified rat liver microsomes oxidized N-nitroso-N-methylvinylamine to form 1,N6-epsilon-Ado in the presence of Ado. Further, 1,N6-epsilon-Ado was also formed in microsomal incubations containing N-nitroso-N-methylethylamine, indicating that desaturation of the ethyl moiety occurs to form a vinyl group and then an epoxide. When NADPH-fortified microsomes were incubated with N-nitroso-N-methylvinylamine, HCHO was formed, and when DNA was included in incubations, 1,N6-epsilon-dAdo and N7-methylGua were isolated from DNA. In the cases of both HCHO and N7-methylGua, product formation was similar to the levels seen with N-nitroso-N,N-dimethylamine and N-nitroso-N-methylethylamine.     

The divergent domains of the NEFA and nucleobindin proteins are derived from an EF-hand ancestor

The human protein NEFA (DNA binding, EF-hand, Acidic region) has previously been isolated from a KM3 cell line and immunolocalized on the plasma membrane, in the cytoplasma, and in the culture medium. Sequence analysis of a cDNA clone encoding NEFA identified a hydrophilic domain, two EF-hands, and a leucine zipper at the C-terminus. These characters are shared with nucleobindin (Nuc). In this paper we have further characterized NEFA and probed its evolutionary origins. Circular dichroism (CD) spectra of recombinant NEFA indicated a helical content of 51% and showed that the EF-hands are capable of binding Ca2+. Experiments with recombinant NEFA and synthesized peptides revealed that the leucine zipper cannot form a homodimer. The leucine zipper may allow heterodimer formation of NEFA and an unknown protein. Phylogenetic analyses suggest that this protein is derived from a four-domain EF-hand ancestor with subsequent duplications and fusions. The leucine zipper and putative DNA-binding domains of NEFA have evolved secondarily from existing EF-hand sequences. These analyses provide insights into how complex proteins may originate and trace the precursor of NEFA to the common ancestor of eukaryotes.     

Possible mechanisms for the nucleation of primary fracture zones during deformation-induced phase transformations in solids: IV. Kinetic mechanisms of submicrocrack nucleation

It is shown that external mechanical actions in condensed matter can cause structural-phase transformations between the states of aggregation, which play a key role in the loss of stability of solids during fracture. These transformations include local melting and sublimation of a solid phase near stress concentrators. This theory is used to develop a kinetic model for the nucleation of primary continuity defects, whose development can result in the appearance of submicrocracks in initially continuous solids.     

Evidence for an inhibition by endogenous galanin of neurogenic cutaneous vasodilatation in the pigeon

The effect of high affinity galanin antagonist M35 on neurogenic cutaneous vasodilatation has been studied in the pigeon using a Laser Doppler Imager. Cutaneous application of mustard oil or antidromic electrical stimulation of a cutaneous nerve produced a small increase in skin blood flow. Close arterial injection of M35 prior to chemical or electrical stimulation resulted in a marked augmentation of the vasodilatory response. This effect was abolished by chronic denervation. The results suggest a nerve-mediated inhibitory effect of endogenous galanin on neurogenic cutaneous vasodilatation in the pigeon skin and provide the first experimental evidence for an inhibitory local regulatory function of cutaneous sensory nerves at least in the avian skin.     

Homogeneous nucleation of liquid from the vapor phase in an expansion cloud chamber

A Wilson expansion cloud chamber has been used to measure the homogeneous vapor-to-liquid nucleation of several materials over the past decade. These data, representing nucleation rate as a function of temperature and supersaturation ratio for toluene, nonane, and water measured in our center, have been re-examined and are presented both in graphical form and as a table of selected data points. The latter is included for ease of comparison with theory. Measurements of the binary homogeneous nucleation of ethanol-water (E-W) mixtures are also presented. The expansion chamber experimental technique and data reduction technique are discussed. This paper is based on a presentation made in the “G. Marshall Pound Memorial Symposium on the Kinetics of Phase Transformations” presented as part of the 1990 fall meeting of TMS, October 8–12, 1990, in Detroit, MI, under the auspices of the ASM/MSD Phase Transformations Committee.     

Elevation of the serum bilirubin diconjugate fraction provides an early marker for cholestasis in the rat

OBJECTIVES: To determine whether the sisters of women with premature ovarian failure (POF) showed a response to gonadotropin stimulation comparable to that of anonymous ovum donors. DESIGN: Historical cohort study. SETTING: Records of 228 consecutive ovum recipients in an academic assisted reproductive technology program. PATIENT(S): Criteria for inclusion were oocyte recipients age < or = 40 years, FSH > 18 mIU/mL (conversion factor to SI unit, 1.00), and/or failure to respond appropriately to controlled ovarian hyperstimulation (COH). Seventy-nine recipients were classified on the basis of whether they received oocytes from anonymous donors (group I, n = 66) or sister donors (group II, n = 13). MAIN OUTCOME MEASURE(S): Controlled ovarian hyperstimulation response, pregnancy rates (PRs), and implantation rates. RESULT(S): The ages of the donors to groups I and II were comparable (31.1 +/- 16.7 versus 29.8 +/- 7.2 years), but those in group II exhibited a higher baseline FSH level (12.8 +/- 2.1 versus 8.6 +/- 5.8 mIU/mL). Group II versus I had a relative risk of 5.1 for cancellation (4 of 13 [30.8%] versus 4 of 66 [6.1%], respectively). In completed cycles of groups I and II, respectively, there was no difference in serum E2 on the day of hCG administration (2,356 +/- 826 versus 1,847 +/- 843 pg/mL; conversion factor to SI unit, 3,671), number of oocytes retrieved (25 +/- 14 versus 22 +/- 13), number of embryos transferred (4.4 +/- 2.1 versus 4.0 +/- 1.0), spontaneous abortion rate (22.7% versus 25.0%), PR (35.5% versus 36.4%), and implantation rate (16.2% versus 16.4%). CONCLUSION(S): There is an increased cancellation rate and, consequently, an overall trend toward decreased ovarian response to gonadotropin stimulation in the sisters of patients with POF. Despite these factors, the implantation rates and PRs of embryos derived from patients reaching retrieval were similar to those from anonymous donors. We recommend counseling women with POF that their sisters may not be ideal ovum donors.     

Dermal uptake of solvents from the vapour phase: an experimental study in humans

An overview of computer models developed since the late seventies, which enable the simulation of the primary effects of spinal cord stimulation (SCS) on nerve fibres, is presented. These models consist of a 3-dimensional volume conductor model, representing anatomical structures and their electrical conductivities, and cable models representing the electrical behaviour of nerve fibres. The characteristics of these models and their relation to anatomy and physiology, as well as the calculation of stimulation-induced electrical fields and their effect on nerve fibre models, are reviewed. It is shown that most characteristics of SCS as predicted by computer modelling correspond well with empirical data. Accordingly, a theoretical framework describing the relations between relevant parameters in SCS is presented. Finally, it is shown how theory and computer modeling are applied to improve the efficacy of SCS by the optimization of its technique, primarily by the design of new epidural electrodes.