Influence of suture tension to the tearing characteristics of the soft tissues: an in vitro experiment

OBJECTIVES: To evaluate the influence of flap tension on the tearing characteristics of mucosal tissue samples in relation to various suture and needle characteristics. MATERIAL AND METHODS: Lining and masticatory mucosal tissue samples obtained from pig jaws were prepared for in vitro testing. Tension tearing diagrams of 60 experiments were traced for 3-0, 5-0 and 7-0 sutures with applied forces up to 20 N. In the second part, the same experiments were repeated with 100 diagrams to test the influence of needle characteristics with 5-0 and 6-0 sutures using only gingival tissue samples. RESULTS: 3-0 sutures mainly lead to tissue breakage at an average of 13.4 N. In contrast, 7-0 sutures only resulted in breakage of the thread at a mean applied force of 3.7 N. With 5-0 sutures, both events occurred at random at a mean force of 14.6 N. Irrespective of the needle characteristics, the mean breaking force for gingival samples with 5-0 and 6-0 sutures was approximately 10 N. CONCLUSIONS: Tissue trauma may be reduced by choosing finer suture diameters, because thinner (6-0, 7-0) sutures lead to thread breakage rather than tissue breakage.

Natural antibodies target virus-antibody complexes to organized lymphoid tissue

Natural antibodies (NA) specific for infectious pathogens are found at low titer (usually <1:40) in the serum of healthy, non-immunized, individuals. Therefore, NA are part of the first line of defence against blood borne microorganisms. They directly neutralize viral infections or lyse pathogens by activating the complement cascade. In addition, recent studies highlighted their role in the pooling of infectious pathogens and other antigens to the spleen. This prevents infection of vital target organs and enhances the induction of adaptive immune responses. Specific T and B-cell responses are exclusively induced in highly organized secondary lymphoid organs including lymph nodes and the spleen. As a consequence, mice with disrupted microorganisation of lymphoid organs have defective adaptive immunity. In addition, some pathogens including lymphocytic choriomeningitis virus (LCMV), Leishmania and HIV developed strategies to destroy the splenic architecture in order to induce an acquired immunosuppression and to establish persistent infection. NA antibodies enhance early neutralizing antibodies in the absence of T help mainly by targeting antigen to the splenic marginal zone. In addition, by activating the complement cascade, NA enhance T cell and T-cell dependent B-cell responses. Therefore, natural antibodies are an important link between innate and adaptive immunity.

The pollen tube: a soft shell with a hard core

Plant cell expansion is controlled by a fine-tuned balance between intracellular turgor pressure, cell wall loosening and cell wall biosynthesis. To understand these processes, it is important to gain in-depth knowledge of cell wall mechanics. Pollen tubes are tip-growing cells that provide an ideal system to study mechanical properties at the single cell level. With the available approaches it was not easy to measure important mechanical parameters of pollen tubes, such as the elasticity of the cell wall. We used a cellular force microscope (CFM) to measure the apparent stiffness of lily pollen tubes. In combination with a mechanical model based on the finite element method (FEM), this allowed us to calculate turgor pressure and cell wall elasticity, which we found to be around 0.3 MPa and 20–90 MPa, respectively. Furthermore, and in contrast to previous reports, we showed that the difference in stiffness between the pollen tube tip and the shank can be explained solely by the geometry of the pollen tube. CFM, in combination with an FEM-based model, provides a powerful method to evaluate important mechanical parameters of single, growing cells. Our findings indicate that the cell wall of growing pollen tubes has mechanical properties similar to rubber. This suggests that a fully turgid pollen tube is a relatively stiff, yet flexible cell that can react very quickly to obstacles or attractants by adjusting the direction of growth on its way through the female transmitting tissue.

Sonication: a valuable technique for diagnosis and treatment of periprosthetic joint infections

BACKGROUND Periprosthetic joint infection (PJI) is the most severe complication, following joint arthroplasty. Identification of the causal microbial factor is of paramount importance for the successful treatment. PURPOSE The aim of this study is to compare the sonication fluid cultures derived from joint prosthetic components with the respective periprosthetic tissue cultures. METHODS Explanted prosthesis components for suspected infection were placed into a tank containing sterile Ringer's solution and sonicated for 1 minute at 40 kHz. Sonication fluid cultures were examined for 10 days, and the number and identity of any colony morphology was recorded. In addition, periprosthetic tissue specimens (>5) were collected and cultured according to standard practice. The duration of antimicrobial interruption interval before culture sampling was recorded. RESULTS Thirty-four patients composed the study group. Sonication fluid cultures were positive in 24 patients (70.5%). Sixteen of thirty four periprosthetic tissue cultures (47.1%) were considered positive, all revealing the same microbial species with the respective sonication fluid cultures: 3 tissue samples showed polymicrobial infection. All tissue cultures were also found positive by the sonication fluid culture. CONCLUSIONS Sonication fluid cultures represent a cheap, easy, accurate, and sensitive diagnostic modality demonstrating increased sensitivity compared to periprosthetic tissue cultures (70.5 versus 47.1%).

Antibodies trap tissue migrating helminth larvae and prevent tissue damage by driving IL-4Rα-independent alternative differentiation of macrophages

Approximately one-third of the world's population suffers from chronic helminth infections with no effective vaccines currently available. Antibodies and alternatively activated macrophages (AAM) form crucial components of protective immunity against challenge infections with intestinal helminths. However, the mechanisms by which antibodies target these large multi-cellular parasites remain obscure. Alternative activation of macrophages during helminth infection has been linked to signaling through the IL-4 receptor alpha chain (IL-4Rα), but the potential effects of antibodies on macrophage differentiation have not been explored. We demonstrate that helminth-specific antibodies induce the rapid trapping of tissue migrating helminth larvae and prevent tissue necrosis following challenge infection with the natural murine parasite Heligmosomoides polygyrus bakeri (Hp). Mice lacking antibodies (JH (-/-)) or activating Fc receptors (FcRγ(-/-)) harbored highly motile larvae, developed extensive tissue damage and accumulated less Arginase-1 expressing macrophages around the larvae. Moreover, Hp-specific antibodies induced FcRγ- and complement-dependent adherence of macrophages to larvae in vitro, resulting in complete larval immobilization. Antibodies together with helminth larvae reprogrammed macrophages to express wound-healing associated genes, including Arginase-1, and the Arginase-1 product L-ornithine directly impaired larval motility. Antibody-induced expression of Arginase-1 in vitro and in vivo occurred independently of IL-4Rα signaling. In summary, we present a novel IL-4Rα-independent mechanism of alternative macrophage activation that is antibody-dependent and which both mediates anti-helminth immunity and prevents tissue disruption caused by migrating larvae.

Identification of the Causative Bacteria in Musculoskeletal Infections Using 16s rDNA - Denaturing Gradient Gel Electrophoresis Analysis

Musculoskeletal infections are infections of the bone and surrounding tissues. They are currently diagnosed based on culture analysis, which is the gold standard for pathogen identification. However, these clinical laboratory methods are frequently inadequate for the identification of the causative agents, because a large percentage (25-50%) of confirmed musculoskeletal infections are false negatives in which no pathogen is identified in culture. My data supports these results. The goal of this project was to use PCR amplification of a portion of the 16S rRNA gene to test an alternative approach for the identification of these pathogens and to assess the diversity of the bacteria involved. The advantages of this alternative method are that it should increase sample sensitivity and the speed of detection. In addition, bacteria that are non-culturable or in low abundance can be detected using this molecular technique. However, a complication of this approach is that the majority of musculoskeletal infections are polymicrobial, which prohibits direct identification from the infected tissue by DNA sequencing of the initial 16S rDNA amplification products. One way to solve this problem is to use denaturing gradient gel electrophoresis (DGGE) to separate the PCR products before DNA sequencing. Denaturing gradient gel electrophoresis (DGGE) separates DNA molecules based on their melting point, which is determined by their DNA sequence. This analytical technique allows a mixture of PCR products of the same length that electrophoreses through agarose gels as one band, to be separated into different bands and then used for DNA sequence analysis. In this way, the DGGE allows for the identification of individual bacterial species in polymicrobial-infected tissue, which is critical for improving clinical outcomes. By combining the 16S rDNA amplification and the DGGE techniques together, an alternative approach for identification has been used. The 16S rRNA gene PCR-DGGE method includes several critical steps: DNA extraction from tissue biopsies, amplification of the bacterial DNA, PCR product separation by DGGE, amplification of the gel-extracted DNA, and DNA sequencing and analysis. Each step of the method was optimized to increase its sensitivity and for rapid detection of the bacteria present in human tissue samples. The limit of detection for the DNA extraction from tissue was at least 20 Staphylococcus aureus cells and the limit of detection for PCR was at least 0.05 pg of template DNA. The conditions for DGGE electrophoreses were optimized by using a double gradient of acrylamide (6 – 10%) and denaturant (30-70%), which increased the separation between distinct PCR products. The use of GelRed (Biotium) improved the DNA visualization in the DGGE gel. To recover the DNA from the DGGE gels the gel slices were excised, shredded in a bead beater, and the DNA was allowed to diffuse into sterile water overnight. The use of primers containing specific linkers allowed the entire amplified PCR product to be sequenced and then analyzed. The optimized 16S rRNA gene PCR-DGGE method was used to analyze 50 tissue biopsy samples chosen randomly from our collection. The results were compared to those of the Memorial Hermann Hospital Clinical Microbiology Laboratory for the same samples. The molecular method was congruent for 10 of the 17 (59%) culture negative tissue samples. In 7 of the 17 (41%) culture negative the molecular method identified a bacterium. The molecular method was congruent with the culture identification for 7 of the 33 (21%) positive cultured tissue samples. However, in 8 of the 33 (24%) the molecular method identified more organisms. In 13 of the 15 (87%) polymicrobial cultured tissue samples the molecular method identified at least one organism that was also identified by culture techniques. Overall, the DGGE analysis of 16S rDNA is an effective method to identify bacteria not identified by culture analysis.

Engineered liposomes sequester bacterial exotoxins and protect from severe invasive infections in mice

Gram-positive bacterial pathogens that secrete cytotoxic pore-forming toxins, such as Staphylococcus aureus and Streptococcus pneumoniae, cause a substantial burden of disease. Inspired by the principles that govern natural toxin-host interactions, we have engineered artificial liposomes that are tailored to effectively compete with host cells for toxin binding. Liposome-bound toxins are unable to lyse mammalian cells in vitro. We use these artificial liposomes as decoy targets to sequester bacterial toxins that are produced during active infection in vivo. Administration of artificial liposomes within 10 h after infection rescues mice from septicemia caused by S. aureus and S. pneumoniae, whereas untreated mice die within 24-33 h. Furthermore, liposomes protect mice against invasive pneumococcal pneumonia. Composed exclusively of naturally occurring lipids, tailored liposomes are not bactericidal and could be used therapeutically either alone or in conjunction with antibiotics to combat bacterial infections and to minimize toxin-induced tissue damage that occurs during bacterial clearance

Detection and quantification of Flavobacterium psychrophilum in water and fish tissue samples by quantitative real time PCR.

BACKGROUND Flavobacterium psychrophilum is the agent of Bacterial Cold Water Disease and Rainbow Trout Fry Syndrome, two diseases leading to high mortality. Pathogen detection is mainly carried out using cultures and more rapid and sensitive methods are needed. RESULTS We describe a qPCR technique based on the single copy gene β' DNA-dependent RNA polymerase (rpoC). Its detection limit was 20 gene copies and the quantification limit 103 gene copies per reaction. Tests on spiked spleens with known concentrations of F. psychrophilum (106 to 101 cells per reaction) showed no cross-reactions between the spleen tissue and the primers and probe. Screening of water samples and spleens from symptomless and infected fishes indicated that the pathogen was already present before the outbreaks, but F. psychrophilum was only quantifiable in spleens from diseased fishes. CONCLUSIONS This qPCR can be used as a highly sensitive and specific method to detect F. psychrophilum in different sample types without the need for culturing. qPCR allows a reliable detection and quantification of F. psychrophilum in samples with low pathogen densities. Quantitative data on F. psychrophilum abundance could be useful to investigate risk factors linked to infections and also as early warning system prior to potential devastating outbreak.

Serological differentiation between Echinococcus granulosus and E. multilocularis infections in man.

An enzyme-linked immunosorbent assay (ELISA) was adapted for the serological differential diagnosis of cystic or alveolar echinococcosis in man caused by Echinococcus granulosus or E. multilocularis respectively. By affinity chromatography using rabbit anti hydatid fluid IgG coupled covalently to CNBr-Sepharose 4B a protein fraction (Em 1) containing shared antigens of both parasites could be isolated from an extract of E. multilocularis metacestode tissue. From the same source another antigen fraction (Em 2) with a high degree of specificity for E. multilocularis was prepared by immunosorption. Antigen Em 1 was equally sensitive for the detection of antibodies against E. granulosus and E. multilocularis, whereas antigen fraction Em 2 appeared to be more specific for E. multilocularis. A correct serological differential diagnosis was achieved in 95% of 57 confirmed cases of human cystic or alveolar echinococcosis by the simultaneous use of both antigen fractions in the ELISA and by comparison of their reactivities.

Multiple infections of rodents with zoonotic pathogens in Austria

Rodents are important reservoirs for a large number of zoonotic pathogens. We examined the occurrence of 11 viral, bacterial, and parasitic agents in rodent populations in Austria, including three different hantaviruses, lymphocytic choriomeningitis virus, orthopox virus, Leptospira spp., Borrelia spp., Rickettsia spp., Bartonella spp., Coxiella burnetii, and Toxoplasma gondii. In 2008, 110 rodents of four species (40 Clethrionomys glareolus, 29 Apodemus flavicollis, 26 Apodemus sylvaticus, and 15 Microtus arvalis) were trapped at two rural sites in Lower Austria. Chest cavity fluid and samples of lung, spleen, kidney, liver, brain, and ear pinna skin were collected. We screened selected tissue samples for hantaviruses, lymphocytic choriomeningitis virus, orthopox viruses, Leptospira, Borrelia, Rickettsia, Bartonella spp., C. burnetii, and T. gondii by RT-PCR/PCR and detected nucleic acids of Tula hantavirus, Leptospira spp., Borrelia afzelii, Rickettsia spp., and different Bartonella species. Serological investigations were performed for hantaviruses, lymphocytic choriomeningitis virus, orthopox viruses, and Rickettsia spp. Here, Dobrava-Belgrade hantavirus-, Tula hantavirus-, lymphocytic choriomeningitis virus-, orthopox virus-, and rickettsia-specific antibodies were demonstrated. Puumala hantavirus, C. burnetii, and T. gondii were neither detected by RT-PCR/PCR nor by serological methods. In addition, multiple infections with up to three pathogens were shown in nine animals of three rodent species from different trapping sites. In conclusion, these results show that rodents in Austria may host multiple zoonotic pathogens. Our observation raises important questions regarding the interactions of different pathogens in the host, the countermeasures of the host's immune system, the impact of the host–pathogen interaction on the fitness of the host, and the spread of infectious agents among wild rodents and from those to other animals or humans.

Characterization of healing tissue in a tooth extraction socket in a rabbit model

The histology of healing in a tooth extraction socket has been described in many studies. The focus of research in bone biology and healing is now centered on molecular events that regulate repair of injured tissue. Rapid progress in cellular and molecular biology has resulted in identification of many signaling molecules (growth factors and cytokines) associated with formation and repair of skeletal tissues. Some of these include members of the transforming growth factor-β superfamily (including the bone morphogenetic proteins), fibroblast growth factors, platelet derived growth factors and insulin like growth factors. ^ Healing of a tooth extraction socket is a complex process involving tissue repair and regeneration. It involves chemotaxis of appropriate cells into the wound, transformation of undifferentiated mesenchymal cells to osteoprogenitor cells, proliferation and differentiation of committed bone forming cells, extracellular matrix synthesis, mineralization of osteoid, maturation and remodeling of bone. Current data suggests that these cellular events are precisely controlled and regulated by specific signaling molecules. A plethora of cytokines; have been identified and studied in the past two decades. Some of these like transforming growth factor beta (TGF-β), vascular endothelial growth factor (VEGF), platelet derived growth factor (PDGF) and fibroblast growth factors (FGFs) are well conserved proteins involved in the initial response to injury and repair in soft and hard tissue. ^ The purpose of this study was to characterize the spatial and temporal localization of TGF-βl, VEGF, PDGF-A, FGF-2 and BMP-2, and secretory IgA in a tooth extraction socket model, and evaluate correlation of spatial and temporal changes of these growth factors to histological events. The results of this study showed positive correlation of histological events to spatial and temporal localization of TGF-β1, BMP-2, FGF-2, PDGF-A, and VEGF in a rabbit tooth extraction model. ^

Activation of target-tissue immune-recognition molecules by double-stranded polynucleotides

Abnormal expression of major histocompatibility complex (MHC) class I and class II in various tissues is associated with autoimmune disease. Autoimmune responses can be triggered by viral infections or tissue injuries. We show that the ability of a virus or a tissue injury to increase MHC gene expression is duplicated by any fragment of double-stranded (ds) DNA or dsRNA introduced into the cytoplasm of nonimmune cells. Activation is sequence-independent, is induced by ds polynucleotides as small as 25 bp in length, and is not duplicated by single-stranded polynucleotides. In addition to causing abnormal MHC expression, the ds nucleic acids increase the expression of genes necessary for antigen processing and presentation: proteasome proteins (e.g., LMP2), transporters of antigen peptides; invariant chain, HLA-DM, and the costimulatory molecule B7.1. The mechanism is different from and additive to that of γ-interferon (γIFN), i.e., ds polynucleotides increase class I much more than class II, whereas γIFN increases class II more than class I. The ds nucleic acids also induce or activate Stat1, Stat3, mitogen-activated protein kinase, NF-κB, the class II transactivator, RFX5, and the IFN regulatory factor 1 differently from γIFN. CpG residues are not responsible for this effect, and the action of the ds polynucleotides could be shown in a variety of cell types in addition to thyrocytes. We suggest that this phenomenon is a plausible mechanism that might explain how viral infection of tissues or tissue injury triggers autoimmune disease; it is potentially relevant to host immune responses induced during gene therapy.

Macroglossia como manifestação de amiloidose : caso clínico

Trabalho Final do Curso de Mestrado Integrado em Medicina, Faculdade de Medicina, Universidade de Lisboa, 2014

Does nuclear tissue infected with bacteria following disc herniations lead to Modic changes in the adjacent vertebrae?

Purpose To investigate the prevalence of infected herniated nucleus material in lumbar disc herniations and to determine if patients with an anaerobic infected disc are more likely to develop Modic change (MC) (bone oedema) in the adjacent vertebrae after the disc herniation. MCs (bone oedema) in vertebrae are observed in 6 % of the general population and in 35-40 % of people with low back pain. These changes are strongly associated with low back pain. There are probably a mechanical cause and an infective cause that causes MC. Several studies on nuclear tissue from herniated discs have demonstrated the presence of low virulent anaerobic microorganisms, predominantly Propionibacterium acnes, in 7-53 % of patients. At the time of a herniation these low virulent anaerobic bacteria may enter the disc and give rise to an insidious infection. Local inflammation in the adjacent bone may be a secondary effect due to cytokine and propionic acid production. Methods Patients undergoing primary surgery at a single spinal level for lumbar disc herniation with an MRI-confirmed lumbar disc herniation, where the annular fibres were penetrated by visible nuclear tissue, had the nucleus material removed. Stringent antiseptic sterile protocols were followed. Results Sixty-one patients were included, mean age 46.4 years (SD 9.7), 27 % female. All patients were immunocompetent. No patient had received a previous epidural steroid injection or undergone previous back surgery. In total, microbiological cultures were positive in 28 (46 %) patients. Anaerobic cultures were positive in 26 (43 %) patients, and of these 4 (7 %) had dual microbial infections, containing both one aerobic and one anaerobic culture. No tissue specimens had more than two types of bacteria identified. Two (3 %) cultures only had aerobic bacteria isolated. In the discs with a nucleus with anaerobic bacteria, 80 % developed new MC in the vertebrae adjacent to the previous disc herniation. In contrast, none of those with aerobic bacteria and only 44 % of patients with negative cultures developed new MC. The association between an anaerobic culture and new MCs is highly statistically significant (P = 0.0038), with an odds ratio of 5.60 (95 % CI 1.51-21.95). Conclusion These findings support the theory that the occurrence of MCs Type 1 in the vertebrae adjacent to a previously herniated disc may be due to oedema surrounding an infected disc. The discs infected with anaerobic bacteria were more likely (P<0.0038) to develop MCs in the adjacent vertebrae than those in which no bacteria were found or those in which aerobic bacteria were found. © Springer-Verlag Berlin Heidelberg 2013.

Caputo Derivatives in Viscoelasticity: A Non-Linear Finite-Deformation Theory for Tissue

Mathematics Subject Classification: 26A33, 74B20, 74D10, 74L15