Immunodominant proteins of Borrelia burgdorferi,the etiological agent of lyme borreliosis

Lyme borreliosis, a multisystem disorder involving the skin, the nervous system, the heart, the joints and many other organs, is a worldwide infectious disease which is transmitted by ticks of the Ixodes complex. Most frequently diagnosis is accomplished by detection of antibodies because the Borrelia are difficult to cultivate. Present serodiagnostic methods, however, are impaired by low sensitivity and unspecific reactions. The selection of immunodominant antigens with low cross-reactivity to other bacteria should improve antibody detection. Borrelia burgdorferi proteins have been analysed for cross-reactivity with immune sera from unrelated bacteria, and sera from patients with different stages of the disease. Suitable antigens for improving serodiagnosis have been detected and are reported here. In view of the immunological heterogeneity of Borrelia proteins, sensitivity of antibody detection may possibly be increased by using recombinant antigens derived from different strains. Immunization with recombinant OspA (a flagellum-associated protein) from a North American isolate protected mice from the challenge with three North American isolates. However, for development of an effective vaccine (especially in Europe), the heterogeneity of OspA has to be considered.This paper was presented at the IUMS Symposium on New Developments in Diagnosis and Control of Infectious Diseases held in conjunction with the Eighth International Congress of Virology, Berlin, Germany, 24–31 August 1990.     

The active species of 'CO2' utilized by reduced ferredoxin:CO2 oxidoreductase from Clostridium pasteurianum.

Reduced ferredoxin:CO2 oxidoreductase (CO2 reductase) from Clostridium pasteurianum catalyzes the reduction of 'CO2' to formate with reduced ferredoxin, an isotopic exchange between 'CO2' and formate in the absence of ferredoxin, and the oxidation of formate to 'CO2' with oxidized ferredoxin. The active species of 'CO2', i.e. CO2 or HCO3 (H2CO3), utilized by the enzyme was determined. The method employed for the species identification was that of Copper et al. (1968). Both 'CO2' reduction to formate and the exchange reaction were studied. Data were obtained which are compatible with those expected if CO2 is the active species. The V and the dissociation constant Ks of the enzyme - CO2 complex in dependence of pH were determined from initial velocity studies of the exchange reaction. V was found to be only slightly affected by pH between 5.5 and 7.5. Ks was markedly dependent on pH; the constant increased with decreasing pH from 0.2 mM at pH 7.5 to 3 mM at pH 5.5.     

Postischaemic circulation disturbances.

Restoration of blood supply after ischaemic conditions in extremities and testes is inhibited by reversible intravasal aggregation of erythrocytes. This process is promoted by the increased permeability of the capillaries associated with the formation of oedema and the entailing increase of the haematocrit. For overcoming the stasis the increased structural viscosity caused by the aggregation of erythrocytes requires an increase in pressure as a starter effect which is not achieved by the flow pressure at once everywhere. Intravenously administered particles of Indian ink mark the formation and dissolution of aggregates. Even areas with originally normal blood supply may be obstructed by the later formation of aggregates. Thrombi on the walls of arterial and venous vessels and other lesions of the intima do not sufficiently explain the disturbance of perfusion. Oedema and extravasating leucocytes are found in the microcirculation. The parenchyma to be supplied shows formation of necrosis.     

Mice lacking the mitochondrial exonuclease MGME1 develop inflammatory kidney disease with glomerular dysfunction

Mitochondrial DNA (mtDNA) maintenance disorders are caused by mutations in ubiquitously expressed nuclear genes and lead to syndromes with variable disease severity and tissue-specific phenotypes. Loss of function mutations in the gene encoding the mitochondrial genome and maintenance exonuclease 1 (MGME1) result in deletions and depletion of mtDNA leading to adult-onset multisystem mitochondrial disease in humans. To better understand the in vivo function of MGME1 and the associated disease pathophysiology, we characterized a Mgme1 mouse knockout model by extensive phenotyping of ageing knockout animals. We show that loss of MGME1 leads to de novo formation of linear deleted mtDNA fragments that are constantly made and degraded. These findings contradict previous proposal that MGME1 is essential for degradation of linear mtDNA fragments and instead support a model where MGME1 has a critical role in completion of mtDNA replication. We report that Mgme1 knockout mice develop a dramatic phenotype as they age and display progressive weight loss, cataract and retinopathy. Surprisingly, aged animals also develop kidney inflammation, glomerular changes and severe chronic progressive nephropathy, consistent with nephrotic syndrome. These findings link the faulty mtDNA synthesis to severe inflammatory disease and thus show that defective mtDNA replication can trigger an immune response that causes age-associated progressive pathology in the kidney.     

Response of rats to the presence of stressed conspecifics as a function of time of day

The physiological response of nonrestrained rats to the presence of immobilized conspecifics during the beginning of the active period and the inactive period was studied. In immobilized animals concentrations of serum corticosterone (SCS), serum glucose, and liver glycogen, and the activity of liver tyrosine aminotransferase (TAT) during both the active and the inactive periods, were consistent with earlier studies. In nonrestrained rats the presence of immobilized conspecifics induced a significant increase in SCS during the active period, whereas it had no effect during the inactive period. The level of TAT was significantly elevated in the nonrestrained rats only during the inactive period and remained unchanged during the active period. The results demonstrate a physiological influence of stressed rats on unstressed conspecifics and provide evidence for regulation of TAT activity that is dependent on the situation and the time of day.     

Conformation, stability, and active-site cysteine titrations of Escherichia coli D26A thioredoxin probed by Raman spectroscopy.

The active-site cysteines (Cys 32 and Cys 35) of Escherichia coli thioredoxin are oxidized to a disulfide bridge when the protein mediates substrate reduction. In reduced thioredoxin, Cys 32 and Cys 35 are characterized by abnormally low pKa values. A conserved side chain, Asp 26, which is sterically accessible to the active site, is also essential to oxidoreductase activity. pKa values governing cysteine thiol-thiolate equilibria in the mutant thioredoxin, D26A, have been determined by direct Raman spectrophotometric measurement of sulfhydryl ionizations. The results indicate that, in D26A thioredoxin, both sulfhydryls titrate with apparent pKa values of 7.5+/-0.2, close to values measured previously for wild-type thioredoxin. Sulfhydryl Raman markers of D26A and wild-type thioredoxin also exhibit similar band shapes, consistent with minimal differences in respective cysteine side-chain conformations and sulfhydryl interactions. The results imply that neither the Cys 32 nor Cys 35 SH donor is hydrogen bonded directly to Asp 26 in the wild-type protein. Additionally, the thioredoxin main-chain conformation is largely conserved with D26A mutation. Conversely, the mutation perturbs Raman bands diagnostic of tryptophan (Trp 28 and Trp 31) orientations and leads to differences in their pH dependencies, implying local conformational differences near the active site. We conclude that, although the carboxyl side chain of Asp 26 neither interacts directly with active-site cysteines nor is responsible for their abnormally low pKa values, the aspartate side chain may play a role in determining the conformation of the enzyme active site.     

Scavenger receptor CD36 mediates uptake of high density lipoproteins in mice and by cultured cells

The mechanisms of HDL-mediated cholesterol transport from peripheral tissues to the liver are incompletely defined. Here the function of scavenger receptor cluster of differentiation 36 (CD36) for HDL uptake by the liver was investigated. CD36 knockout (KO) mice, which were the model, have a 37% increase (P = 0.008) of plasma HDL cholesterol compared with wild-type (WT) littermates. To explore the mechanism of this increase, HDL metabolism was investigated with HDL radiolabeled in the apolipoprotein (¹²⁵I) and cholesteryl ester (CE, [³H]) moiety. Liver uptake of [³H] and ¹²⁵I from HDL decreased in CD36 KO mice and the difference, i. e. hepatic selective CE uptake ([³H]¹²⁵I), declined (–33%, P = 0.0003) in CD36 KO compared with WT mice. Hepatic HDL holo-particle uptake (¹²⁵I) decreased (–29%, P = 0.0038) in CD36 KO mice. In vitro, uptake of ¹²⁵I-/[³H]HDL by primary liver cells from WT or CD36 KO mice revealed a diminished HDL uptake in CD36-deficient hepatocytes. Adenovirus-mediated expression of CD36 in cells induced an increase in selective CE uptake from HDL and a stimulation of holo-particle internalization. In conclusion, CD36 plays a role in HDL uptake in mice and by cultured cells. A physiologic function of CD36 in HDL metabolism in vivo is suggested.