Crystalline and disordered state of poly(dihexylsilylene) copolymers

A systematic comparison of random copolymers, derived from poly(dihexylsilylene) (PDHS) by incorporation of monomeric units with shorter unbranched alkyl side chains, has been carried out based on calorimetry, variable temperature UV spectroscopy, and ²⁹Si MAS (magic angle spinning) solid state NMR investigations. Also, hexylmethylsilylene units and branched monomers have been copolymerized. Up to 10% comonomer with shorter linear side chains (i. e., pentyl to propyl) could be incorporated into PDHS without impeding the all-trans order of the crystalline phase. In this case, the UV absorption maximum of the crystalline low-temperature phase was affected only slightly according to the length and fraction of the comonomer side chains. A less ordered crystal structure (λ_max = 345–355 nm) was observed when the content of comonomers with shorter side chains was about 20%. Yet, all these materials form conformationally disordered mesophases. A clear disordering transition and corresponding thermochromism was not observed any more when 50% of propyl side chains were incorporated. The order of the crystalline and the mesophase is also strongly perturbed if only a small fraction (4%) of the side chains are branched at C².     

The relationship between the electromyogram-amplitude and isometric extension torques of neck muscles at different positions of the cervical spine

A group of 12 healthy men volunteered for the experiment. Electromyograms (EMG) were obtained from semispinalis capitis, splenius capitis, levator scapulae, and trapezius muscles. The flexion angle of the cervical spine was precisely adjusted to 0°, 10°, 20°, and 30° relative to the horizontal, with a constant angle of the atlanto-occipital joint. The subjects made eight short (about 2 s) vertical extension forces (6%, 12%,18%, 24%, 30%, 36%, 42%, and 48% of maximal voluntary peak contraction force). For each position, the centre of pressure under the head was determine as the basis for the calculation of the external lever arm. The presence of motor endplate regions was ascertained by multiple surface electrodes. The slopes of individual linear regression lines for the root mean square (rms)-values were dependent on the existence of endplates in the area of the electrodes — endplates caused smaller rms values per Newton metres of external torque. Significant intersubject differences between regression equations could not be eliminated by the normalization of EMG-parameters and/or torques. The elimination of gravity, the continuous monitoring of positions, and the consideration of localization of motor endplate regions were essential prerequisites for the acquisition of reliable relationships between EMG of different neck muscles and external torques. Two important conclusions were derived for the prediction of torques from EMG measurements: firstly, individual regression equations which take into account the position of the head and neck should be used; secondly, normalization procedures do not justify the application of average regressions to a group of subjects.     

In vitro study on the impact of fish sera on the survival and fine structure of the eel-pathogenic acanthocephalanParatenuisentis ambiguus

The effects of fish sera on the growth and fine structure of infective larvae of the eel-pathogenic acanthocephalanParatenuisentis ambiguus (Eoacanthocephala: Tenuisentidae) were studied under in vitro conditions using sera from the final hostAnguilla anguilla and from two accidental fish hosts as well as fetal calf serum. As controls larvae were also kept in medium in the absence of serum and in experimentally infected eels. Sera from the accidental fish hosts carp and rainbow trout exerted toxic effects on the acanthocephalans. Worms maintained in medium containing sera from these two fish were contracted and displayed inverted probosces. Moreover, the tegument exhibited vacuolization and the formation of necrotic areas, including lysis of the mitochondria. Due to these effects, the parasites died at 21 (rainbow trout) or 21–50 days (carp) postincubation. Eel sera had no toxic effect on the infective larvae. The growth of the larvae in medium depended on the composition of the latter, but was reduced as compared with that in the natural final host. Based on these results, we conclude that components of the hosts' blood sera play a role in the determination of the host specificity ofP. ambiguus.     

Dioxygen sensitivity of a photoexcited thin film of phthalocyanine dispersed in poly(N-vinylcarbazole)

Thin films (thickness 0,3 μm) of a mixture of phthalocyaninatozinc(II) ( 1 ) and poly(1-vinylcarbazole) ( 2 ) are prepared by coating from a solution on ITO glass. The blue coloured films are investigated in an electrochemical cell under irradiation with visible light. In the presence of dioxygen from air the films exhibit an intensive cathodic photocurrent. The action spectrum of the photocurrent corresponds to the absorption spectrum of the phthalocyanine. The results show a device which can be switched by air and visible light.     

Fibre type specific expression of Leu19-antigen and N-CAM in skeletal muscle in various stages after experimental denervation

Leu-19 antigen, which seems to be identical with neural cell adhesion molecule (N-CAM), plays a major role in the innervation of muscle cells, and in adult muscle appears after denervation and during regeneration of muscle fibres, where it acts as part of a signalling system increasing the probability of re-innervation. This combined enzyme-histochemical and immunohistochemical study examined whether this signalling process was regulated in a uniform or differential pattern for type 1 and type 2 muscle fibres. The subscapular nerve of 18 rabbits was transsected with subsequent complete denervation of the supraspinatus muscle. Leu19 and N-CAM immunohistochemistry was performed 2 to 64 days after surgery. Whereas in normal muscle there are virtually no Leu-19/N-CAM positive muscle fibres; from day 2 after denervation an increasing proportion of fibres expressed Leu-19/N-CAM, prior to any neurogenic atrophy. In the early stage of denervation Leul9/N-CAM expression was confined to type 1 fibres. After 11 days nearly all fibres were Leul9/N-CAM positive irrespective of their fibre type. Sixty-four days after denervation type 1 fibres became Leu19/N-CAM negative, while atrophic type 2 fibres showed intensive staining. Thus, expression of Leu-19 antigenity is differently regulated in both fibre types.     

Immunization of Aotus Monkeys with Recombinant Plasmodium falciparum Hybrid Proteins Does Not Reproducibly Result in Protection from Malaria Infection

Plasmodium falciparum antigens SERP, HRPII, MSAI, and 41-3 have shown promise as vaccine components. This study aimed at reproducing and extending previous results using three hybrid molecules. Antibody responses were reproduced in Aotus monkeys, but solid protection from a P. falciparum blood-stage challenge that showed an unintendedly enhanced pathogenicity was not observed.The increasing drug resistance of Plasmodium falciparum, the most pathogenic human malaria parasite, underlines the need for an effective malaria vaccine. Identification, testing, and optimization of candidate molecules originating from all developmental stages of the parasite are under way. Previously, a successful trial in Aotus monkeys employed the Escherichia coli-expressed hybrid proteins MS2/SERP/HRPII and SERP/MSAI/HRPII (11). Both hybrid proteins contain a region of the serine repeat protein SERP (1, 9), including two putative T-cell epitopes (13) and previously shown to induce a partial protective response in Aotus monkeys (5), and the C-terminal half of the histidine-rich protein HRPII (14), which has also been shown to induce a partially protective response (5, 8). SERP/MSAI/HRPII contains in addition a conserved N-terminal region of the merozoite surface antigen MSAI (7) that includes at least four T-cell epitopes (3, 6). Here we report on further analysis of three hybrid proteins of this type in a vaccination trial with Aotus monkeys. Two of the proteins, SERP/HRPII and SERP/MSAI/HRPII, are improved versions of the hybrid proteins mentioned above, obtained by deleting nonmalaria protein regions and changing an internal restart residue (methionine-729 of SERP) into alanine. Thus, the SERP/HRPII hybrid protein comprises residues 630 to 893 of SERP fused to the 189 C-terminal residues of HRPII, and SERP/MSAI/ HRPII comprises residues 630 to 764 of SERP fused to residues 146 to 259 of MSAI, which is fused to the 189 C-terminal residues of HRPII. SERP/41-3/HRPII contains the same components as SERP/HRPII, and additionally includes residues 77 to 188 of antigen 41-3 (10), which was previously shown to confer protection against a P. falciparum challenge (5). The internal restart residue (methionine-100) was also mutagenized into alanine and another residue, arginine-319, was changed into leucine to prevent proteolytic degradation. SERP/MSAI/HRPII was partially purified to a final purity of about 30%, as described previously (8), in order to match the quality of the proteins used in the successful previous trials (5, 11). The other two hybrid proteins were purified from bacterial lysates to over 90% purity by size exclusion chromatography (SERP/41-3/HRPII) or by sequential cation and anion exchange and then size exclusion chromatography (SERP/ HRPII) (data not shown). The final products were dialyzed against phosphate-buffered saline–3 M urea and adjusted to 100 μg of protein per ml. Efficacy was tested following an experimental protocol identical to the one used in the previous successful trial (11).Fifteen laboratory-raised Aotus azarae boliviensis karyotype VI monkeys were randomly assigned to one of four experimental groups (three groups of four and one group of three monkeys) and immunized with 1 ml of antigen or with the diluent alone (control group), both mixed with 100 μl of polyalphaolefin (4) as an adjuvant, on days 0, 21, and 42. Each vaccine dose was administered subcutaneously at two separate sites in the right and left flank and was well tolerated. The seroconversion results, as measured by enzyme-linked immunosorbent assay with SERP/HRPII as the solid-phase antigen and peroxidase-labelled rabbit anti-human immunoglobulin G (1:10,000 dilution; Pierce) as the secondary antibody, are shown in Fig. ​Fig.1.1. All experimental monkeys developed comparable antibody responses to SERP/HRPII, irrespective of the immunogen. Control monkey sera did not react significantly (not shown). A boosting effect is obvious after the second injection in all three groups (Fig. ​(Fig.1),1), as well as after the third SERP/41-3/HRPII injection (Fig. ​(Fig.1C).1C). This is similar to the seroconversion pattern observed previously (11). Prechallenge sera were also tested by immunofluorescence (IFA) for reactivity with P. falciparum schizonts. All preimmune sera and control group immune sera were negative (1:100 dilution). IFA titers from the experimental animals were all 1:1,600, except for animals A381 and A462 (titer, 1:800) and A452 and A292 (titer, 1:3,200). Thus, antibodies specific for native parasite determinants were induced. The relatively low IFA titers were comparable to those obtained in previous successful trials (5, 11). Open in a separate windowFIG. 1Development of antibody responses in Aotus monkeys during the immunization period as determined by enzyme-linked immunosorbent assay. Monkeys in different immunization groups were immunized at weeks 0, 3, and 6 (indicated by arrows) and challenged at week 8 (indicated by an asterisk). All sera were tested for reactivity with SERP/HRPII in a 1:100 dilution. The hybrid antigens used for immunization were SERP/MSAI/HRPII (A), SERP/HRPII (B), and SERP/41-3/HRPII (C). Sera of the three control monkeys remained negative in this assay (not shown). OD, optical density.At week 7 all monkeys were splenectomized, and at week 8 they received intravenously 2 × 10⁶ parasitized erythrocytes, which had been isolated from an Aotus monkey infected with an in vivo-passaged FUP-Cayenne isolate of P. falciparum (a kind gift of W. E. Collins) (Fig. ​(Fig.1).1). Monkey A293 appeared to have no spleen, although there was no prior history of splenectomy. The immunoglobulin G response of monkey A293 was nevertheless comparable to that of the other animals (Fig. ​(Fig.1B).1B). Figure ​Figure22 shows the course of parasitemia after challenge. Two of three control animals rapidly developed a parasitemia which required mefloquine therapy (20 mg/kg of body weight orally) when parasitemia reached 10% (day 8 for A371 and day 10 for A320). In A432, parasitemia developed to 8.6% (day 10) and then fluctuated until rapidly reaching 19% (day 21), at which point mefloquine was administered (Fig. ​(Fig.2A).2A). None of the three immunized groups showed a solid protective response (Fig. ​(Fig.2B2B to D). A340 (SERP/HRPII group) (Fig. ​(Fig.2C)2C) and A292 (SERP/41-3/HRPII group) (Fig. ​(Fig.2D)2D) showed low fluctuating parasitemias with a peak around 2.5% at the end of the observation period (day 25). Otherwise, parasitemias of the experimental animals did not significantly differ from those of the controls. No obvious correlation between prechallenge antibody levels and protection was evident. Open in a separate windowFIG. 2Course of infection with the FUP-Cayenne isolate of P. falciparum in control Aotus monkeys (A) and in Aotus monkeys immunized with SERP/MSAI/HRPII (B), SERP/HRPII (C), and SERP/41-3/HRPII (D). Parasitemias of ≥10% were cured with mefloquine.It seems unlikely that small conservative changes designed to improve SERP/HRPII and SERP/MSAI/HRPII expression in E. coli and to remove nonrelevant sequences adversely affected immune response development. After challenge, parasitemia developed markedly faster than in the previous trial, which had shown protection. Challenge with 2 × 10⁶ parasitized erythrocytes now resulted in high parasitemias on days 7 to 9 in 2 of the 3 controls and in 6 of the 12 experimental animals, whereas previously controls were untreated until day 14 (11). Also, one control and three experimental animals suffered recrudescence, which was not seen previously (11) or with later infections with the same parasite stock (5). It is remarkable that this apparent enhanced pathogenicity developed after a single passage in A. nancymai just before the present trial started. It is likely that this unintended pathogenicity influenced the experimental outcome. The protection of two monkeys in the SERP/HRPII and SERP/41-3/HRPII group may, however, reveal some protective effect of these vaccine candidates.For demonstration of the protective potential of antigens in the primate model the pathogenicity of the challenge strain in the respective primate (sub)species, i.e., the equilibrium of immune response and pathogenicity, seems to be crucial (2, 12). The disturbance of this equilibrium may explain the discrepancy between previous successful trials (5, 11) and the present study. Recombinant proteins shown to be protective in the Aotus model (5, 11) failed to protect Saimiri monkeys, in which the course of parasitemia is quite different from that observed in Aotus monkeys. Similarly, no protection could be demonstrated in A. nancymai against the same challenge strain as was used in the successful trials with A. azarae boliviensis and A. lemurinus griseimembra (7a). The poor standardization of these models due to the scarcity of monkeys susceptible to human malaria remains an obstacle for the evaluation of human malaria vaccine candidates.     

Heterogeneity of Borrelia burgdorferi sensu lato demonstrated by an ospA-type-specific PCR in synovial fluid from patients with Lyme arthritis

Borrelia burgdorferi sensu lato, the etiological agent of Lyme borreliosis, has been divided into three genospecies: B. burgdorferi sensu stricto (OspA-type 1), B. afzelii (OspA-type 2) and B. garinii (OspA-type 3–7). Whereas in Europe B. afzelii (OspA-type 2) is predominant among human skin isolates and B. garinii (OspA-type 3–7) among human CSF isolates, some previous serological studies suggested that Lyme arthritis is also associated with B. burgdorferi sensu stricto in Europe. In the present study we designed ospA type-specific PCRs and identified four different ospA types associated with Lyme arthritis. Our study group consisted of 20 patients with positive serology (ELISA and immunoblotting) and clinical criteria for Lyme arthritis. B. burgdorferi DNA was detected in 13 patients and in none of 10 control patients from synovial fluid. We identified ospA-type 1 (26.6%), ospA-type 2 (33.3%), ospA-type 4 (6.6%) and ospA-type 5 (33.3%). Our conclusion is that in Europe B. burgdorferi sensu lato strains causing Lyme arthritis are considerably heterogeneous and that there is no prevalence of certain genospecies or OspA-types among this strains. Received: 14 May 1998     

Eccentric visual acuity in patients with macular disease

A series of cards each containing a two dimensional array of identical Snellen "E's" was used to determine best eccentric visual acuity in patients with macular disease having Snellen visual acuity of 20/70 or worse. Each "full field E" card simultaneously presents the same letter to foveal and parafoveal areas. This test can therefore determine quickly if potentially useful vision is present in any area of the central visual field. In our study of 37 eyes, 70% demonstrated potential visual acuity at least two times better than visual acuity measured by conventional methods, and 20% demonstrated at least a fourfold improvement. This suggests that most patients with macular disease do not spontaneously employ their best remaining area of retina for fixation.     

Demonstration of Ri-type adenosine receptors in bovine myocardium by radioligand binding

Summary Adenosine has been shown to have negative inotropic, chronotropic and dromotropic effects on the heart. The pharmacological profiles of these effects suggest that they are mediated via R_i (A₁) adenosine receptors, but a direct demonstration of these receptors is still missing. In the present study we report direct labelling of these receptors with (-)N⁶-[¹²⁵I]-p-hydroxyphenylisopropyladenosine ([¹²⁵I]HPIA). The radioligand bound in a saturable and reversible manner to a crude membrane preparation, the B _max-value was 30.5 fmol/mg protein and the K _D-value 1.1 nmol/l. A similar affinity of the ligand was obtained in kinetic and competition experiments. Competition experiments with a variety of adenosine analogues gave a pharmacological profile characteristic of R_i adenosine receptors with high affinities of N⁶-substituted derivatives and a marked stereospecificity for N⁶-phenylisopropyladenosine (PIA). Purification of the membrane preparation by density gradient centrifugation resulted in a 30-fold increase in the number of binding sites which was paralleled by a similar increase in the number of binding sites for [³H]ouabain. Guanine nucleotides decreased binding of [¹²⁵I]HPIA in a dose-dependent manner, but the IC₅₀-values were considerably higher than those reported in other tissues. Finally, binding of [¹²⁵I]HPIA appeared to be entropy-driven which has been shown to be characteristic of agonist binding to R_i adenosine receptors. These results suggest the presence of R_i adenosine receptors in ventricular myocardium which may be responsible for the mediation of the effects of adenosine and its analogues.Abbreviations [¹²⁵I]HPIA (-)N⁶-[¹²⁵I]-p-hydroxyphenylisopropyladenosine - (-)IHPIA (-)N⁶-iodo-p-hydroxyphenylisopropyladenosine - (+)/(-)PIA (+)/(-)N⁶-phenylisopropyladenosine - CHA N⁶-cyclohexyladenosine - NECA 5-N-ethylcarboxamidoadenosine - App(NH)p 5-adenylylimidodiphosphate - Gpp(NH)p 5-guanylylimidodiphosphate