The Role of DbpA and B adhesins in the pathogenesis of Lyme borreliosis

Lyme borreliosis is a tick-borne infectious disease common in the Northern hemisphere. It is most commonly caused by B. burgdorferi sensu stricto, B. garinii or B. afzelii. The Borrelia bacteria have several surface proteins, or adhesins, which mediate attachment to different tissues and molecules in the host. Two of these adhesins, Decorin binding protein A and B (DbpA and B) facilitate bacterial adhesion to extracellular proteoglycan decorin. The aim, therefore, of this study is to characterize and compare the biological activities of DbpA and B of the three genospecies and elucidate the role of these interactions in the pathogenesis of Lyme borreliosis. This study shows that Dbps have different binding properties to decorin and decorin expressing cells. In addition to decorin, the Dbps facilitate bacterial adhesion to another proteoglycan, biglycan, and biglycan expressing cells. Biglycan as a molecule is highly similar to decorin, but its tissue distributions in humans and other mammals vary significantly. Borrelia is able to attach to human vascular endothelial cells through an interaction with biglycan, and the Dbps of B. garinii facilitate the flow tolerant bacterial adhesion to endothelial cells. Furthermore, DbpA and B adhesins are both needed for full arthritis development in mice, and they enable the bacteria to persist as DNA after the antibiotic treatment in mice. This study demonstrates the biological differences of DbpA and B of B. burgdorferi sensu stricto, B. garinii and B. afzelii, and reveals a greater role of the adhesins in the pathogenesis of Lyme borreliosis than was previously assumed.DbpA- ja B-adhesiiniproteiinien merkitys Lymen borrelioosin patogeneesissä Lymen borrelioosi on Ixodes-puutiaisten (kansankielellä punkkien) välityksellä leviävä infektiotauti, jota tavataan pohjoisella pallonpuoliskolla etenkin Pohjois- ja Keski-Euroopassa sekä Pohjois-Amerikassa. Viimeisen vuosikymmenen aikana taudin levinneisyysalue on laajentunut merkittävästi rannikkoalueilta Etelä- ja Keski-Suomeen. Ihmiselle tautia aiheuttaa pääasiassa kolme eri Borrelia burgdorferi sensulato -alalajia: Borrelia burgdorferi sensu stricto, Borrelia garinii ja Borrelia azfelii, joiden kudoshakuisuus ja taudinkuva eroavat toisistaan. Molekyylitason mekanismeja, jotka selittäisivät Borrelia-alalajien erilaisen käyttäytymisen ihmisessä, ei vielä täysin tunneta. Borrelia-bakteerin kyky sitoutua soluväliaineen dekoriiniin DbpA- ja DbpB-adhesiiniproteiinien (engl. Decorin binding protein) välityksellä on osoitettu tärkeäksi infektion kannalta. Dekoriinia ilmennetään sidekudoksissa, erityisesti ihossa ja nivelissä. Tutkimuksen tavoitteena oli kuvata eri Borrelia-alalajien Dbp-proteiinien sitoutuminen dekoriiniin ja muihin mahdollisiin ligandeihin, sekä selvittää tämän sitoutumisaktiivisuuden merkitys Lymen borrelioosin patogeneesissä. Tutkimuksessa havaittiin eri Borrelia-alalajien Dbp-proteiineilla olevan erilainen kyky sitoutua dekoriiniin ja dekoriinia ilmentäviin soluihin. Lisäksi Dbp-proteiinien havaittiin sitoutuvaan biglykaani-nimiseen molekyyliin, ja sitä ilmentäviin soluihin. Biglykaani on dekoriinin kaltainen proteoglykaani, mutta sen kudosjakauma elimistössä eroaa merkittävästi dekoriinin jakaumasta. Borrelia-bakteerien osoitettiin sitoutuvan verisuonten endoteelisoluihin biglykaanin välityksellä. Keskeinen havainto oli B. gariniin-alalajin DbpA- ja B-proteiinien merkitys endoteelisitoutumisessa myös virtauksen alla verenkiertoa simuloivassa koeasetemassa. Tutkimuksessa havaittiin Dbp-proteiineilla olevan tärkeä merkitys myös bakteerin leviämisessä, nivelturvotuksen synnyssä ja antibioottihoidon jälkeisessä DNA-persistoinnissa Lymen borrelioosin hiirimallissa. Nämä tutkimustulokset tuovat merkittävästi lisää tietoa eri Borrelia-alalajien Dbp-proteiinien sitoutumisominaisuuksista, sekä näiden sitoutumisominaisuuksien merkityksestä Borrelia-infektion leviämisessä, niveltulehduksen synnyssä ja antibioottihoidon jälkeisessä bakteeri-DNA:n persistoinnissa.Siirretty Doriast

Nanomechanical mechanisms of Lyme disease spirochete motility enhancement in extracellular matrix

As opposed to pathogens passively circulating in the body fluids of their host, pathogenic species within the Spirochetes phylum are able to actively coordinate their movement in the host to cause systemic infections. Based on the unique morphology and high motility of spirochetes, we hypothesized that their surface adhesive molecules might be suitably adapted to aid in their dissemination strategies. Designing a system that mimics natural environmental signals, which many spirochetes face during their infectious cycle, we observed that a subset of their surface proteins, particularly Decorin binding protein (Dbp) A/B, can strongly enhance the motility of spirochetes in the extracellular matrix of the host. Using single-molecule force spectroscopy, we disentangled the mechanistic details of DbpA/B and decorin/laminin interactions. Our results show that spirochetes are able to leverage a wide variety of adhesion strategies through force-tuning transient molecular binding to extracellular matrix components, which concertedly enhance spirochetal dissemination through the host. Martin Strnad, Yoo Jin Oh, and colleagues use single-molecule force spectroscopy and an extracellular matrix (ECM) analog that mimics natural tick feeding to show that the surface proteins DbpA/B can enhance spirochete motility in the ECM of the host. These results show that spirochetes can tune their transient molecular binding to ECM components to enhance spirochetal dissemination through the host

Conserved lysine residues in decorin binding proteins of Borrelia garinii are critical in adhesion to human brain microvascular endothelial cells

Lyme borreliosis is a tick-borne disease caused by Borrelia burgdorferi sensu lato spirochetes (Lyme borreliae). When the disease affects the central nervous system, it is referred to as neuroborreliosis. In Europe, neuroborreliosis is most often caused by Borrelia garinii. Although it is known that in the host Lyme borreliae spread from the tick bite site to distant tissues via the blood vasculature, the adherence of Lyme borreliae to human brain microvascular endothelial cells has not been studied before. Decorin binding proteins are adhesins expressed on Lyme borreliae. They mediate the adhesion of Lyme borreliae to decorin and biglycan, and the lysine residues located in the binding site of decorin binding proteins are important to the binding activity. In this study, we show that lysine residues located in the canonical binding site can also be found in decorin binding proteins of Borrelia garinii, and that these lysines contribute to biglycan and decorin binding. Most importantly, we show that the lysine residues are crucial for the binding of Lyme borreliae to decorin and biglycan expressing human brain microvascular endothelial cells, which in turn suggests that they are involved in the pathogenesis of neuroborreliosis.</p

Preperitoneal Fat Grafting Inhibits the Formation of Intra-abdominal Adhesions in Mice

BACKGROUND: Adhesion formation contributes to postoperative complications in abdominal and gynaecological surgery. Thus far, the prevention and treatment strategies have focused on mechanical barriers in solid and liquid form, but these methods are not in routine use. As autologous fat grafting has become popular in treatment of hypertrophic scars because of its immunomodulatory effects, we postulated that fat grafting could also prevent peritoneal adhesion through similar mechanisms.METHODS: This was a control versus intervention study to evaluate the effect of fat grafting in the prevention on peritoneal adhesion formation. An experimental mouse model for moderate and extensive peritoneal adhesions was used (n = 4-6 mice/group). Adhesions were induced mechanically, and a free epididymal fat graft from wild type or CAG-DsRed mice was injected preperitoneally immediately after adhesion induction. PET/CT imaging and scaling of the adhesions were performed, and samples were taken for further analysis at 7 and 30 days postoperation. Macrophage phenotyping was further performed from peritoneal lavage samples, and the expression of inflammatory cytokines and mesothelial layer recovery were analysed from peritoneal tissue samples.RESULTS: Fat grafting significantly inhibited the formation of adhesions. PET/CT results did not show prolonged inflammation in any of the groups. While the expression of anti-inflammatory and anti-fibrotic IL-10 was significantly increased in the peritoneum of the fat graft-treated group at 7 days, tissue-resident and repairing M2 macrophages could no longer be detected in the fat graft at this time point. The percentage of the continuous, healed peritoneum as shown by Keratin 8 staining was greater in the fat graft-treated group after 7 days.CONCLUSIONS: Fat grafting can inhibit the formation of peritoneal adhesions in mice. Our results suggest that fat grafting promotes the peritoneal healing process in a paracrine manner thereby enabling rapid regeneration of the peritoneal mesothelial cell layer.</div

<i>B</i>. <i>burgdorferi</i> culture and PCR results of Experiment IV at 15 weeks of infection.

<p>ND Not determined</p><p><i>B</i>. <i>burgdorferi</i> culture and PCR results of Experiment IV at 15 weeks of infection.</p

<i>B</i>. <i>burgdorferi</i> culture and PCR results of Experiment III at two weeks of infection.

<p><i>B</i>. <i>burgdorferi</i> culture and PCR results of Experiment III at two weeks of infection.</p

<i>B</i>. <i>burgdorferi</i> PCR results of Experiment II at 15 weeks of infection.

<p>a One sample not available for <i>flaB</i> PCR</p><p><i>B</i>. <i>burgdorferi</i> PCR results of Experiment II at 15 weeks of infection.</p

IgG antibody levels in mouse serum samples.

<p>Antibody levels were measured using enzyme immunoassays with whole <i>B</i>. <i>burgdorferi</i> lysate as antigen. In panel A, the results are from Experiment II, in panel B from Experiment IV, and in panels C and D combined from Experiments II and III. The results in each panel are obtained from an individual analysis. Each symbol represents the result of one animal. Results are expressed as OD492 values and all samples were analysed in duplicate. The line indicates the mean of each group. Groups with the same letter do not differ at 5% level of probability (Tukey’s HSD test, panels A and B). * P ≤ 0.05, *** P ≤ 0.001.</p

Joint swelling and histology.

<p>In experiment I (A), II (B) and IV (C), the development of joint swelling was monitored by measuring the medio-lateral diameter of the hind tibiotarsal joints once a week. Each curve represents the mean of the study group. Asterisks denote significant difference from uninfected mice (P ≤ 0.05). Paraffin-embedded tissue sections were prepared from Δ<i>dbpAB</i>/<i>dbpAB</i> (D; group 7) and Δ<i>dbpAB</i> (E; group 8) infected and uninfected control (F; group 6) mice at 15 weeks of infection and stained for HE. Arrowheads indicate the synovial membrane, and asterisks indicate articular cartilage surface in panels D and E. The original magnification in panels D and E is ×400 and in panel <i>F</i> ×20. Panel F indicates the anatomical structure shown in panels D and E. “Cef” Ceftriaxone treatment, “aTα” anti-TNF-alpha treatment.</p

Design of the mouse experiments.

<p>In Experiment I, four Δ<i>dbpAB</i>/<i>dbpAB</i> (group 2), eight Δ<i>dbpAB</i>/<i>dbpA</i> (group 3), eight Δ<i>dbpAB</i>/<i>dbpB</i> (group 4), two Δ<i>dbpAB</i> (group 5) infected animals and two uninfected control animals (group 1) were killed at seven weeks of infection. In Experiment II, 16 infected animals (groups 4 and 5) were treated with ceftriaxone and 16 (groups 6 and 7) with ceftriaxone and anti-TNF-alpha. The ceftriaxone treatment was started at two weeks (25 mg/kg twice a day for 5 days) and the anti-TNF-alpha treatment at seven weeks of infection (10 mg/kg once a week for 4 weeks). Ear biopsy samples were collected at 6 and 9 weeks of infection to monitor the dissemination of the infection. In Experiment III, mice were killed at two weeks to study infection kinetics and bacterial load in joints. In Experiment IV, eight infected animals were treated with ceftriaxone at six weeks of infection (groups 14 and 15).</p