An investigation of herpes simplex virus type 1 latency in a novel mouse dorsal root ganglion model suggests a role for ICP34.5 in reactivation.

After a primary lytic infection at the epithelia, herpes simplex virus type 1 (HSV-1) enters the innervating sensory neurons and translocates to the nucleus, where it establishes a quiescent latent infection. Periodically, the virus can reactivate and the progeny viruses spread back to the epithelium. Here, we introduce an embryonic mouse dorsal root ganglion (DRG) culture system, which can be used to study the mechanisms that control the establishment, maintenance and reactivation from latency. Use of acyclovir is not necessary in our model. We examined different phases of the HSV-1 life cycle in DRG neurons, and showed that WT HSV-1 could establish both lytic and latent form of infection in the cells. After reactivating stimulus, the WT viruses showed all markers of true reactivation. In addition, we showed that deletion of the γ(1)34.5 gene rendered the virus incapable of reactivation, even though the virus was clearly able to replicate and persist in a quiescent form in the DRG neurons.

Mechanical properties and in vivo performance of load-bearing fiber-reinforced composite intramedullary nails with improved torsional strength.

Fiber-reinforced composites (FRC) could be feasible materials for fracture fixation devices if the mechanical properties of the composites are congruent with the local structural properties of bone. In a recently developed FRC implant, bisphenol A dimethacrylate (BisGMA) and triethylene glycol dimethacrylate (TEGDMA) resin was reinforced with unidirectional E-glass fibers. The addition of a braided glass fiber sleeving to the unidirectional fibers increased the torsional strength (99.5MPa) of the FRC implants at the expense of the flexural strength (602.0MPa). The flexural modulus was 15.3GPa. Two types of FRC intramedullary nails were prepared; first type was FRC as such, second type was FRC with a surface layer of bioactive glass (BG) granules. Experimental oblong subtrochanteric defect was created in 14 rabbits. The defect, which reduced the torsional strength of the bones by 66%, was fixed with an FRC intramedullary nail of either type. The contralateral intact femur served as the control. This model simulated surgical stabilization of bone metastasis. After 12 weeks of follow-up, the femurs were harvested and analyzed by torsional testing, micro-CT and hard tissue histology. Healed undisplaced peri-implant fractures were noticed in half of the animals irrespective of the type of FRC implant. Torsional testing showed no significant differences between the implantation groups. The torsional strength of the bones stabilized by either type of FRC implant was 83% of that of the contralateral femurs. In histological analysis, no implant debris and no adverse tissue reactions were observed. While the mechanical properties of the modified FRCs were suboptimal, the FRC intramedullary nails supported the femurs without structural failure, even in the cases of peri-implant fractures.

Toll-like receptor 9 ligands enhance mesenchymal stem cell invasion and expression of matrix metalloprotease-13.

Human mesenchymal stem cells (hMSCs) are multipotent cells that are found in the bone marrow. Inflammation and tissue damage mobilize MSCs and induce their migration towards the damaged site through mechanisms that are not well defined. Toll-like receptor-9 (TLR9) is a cellular receptor for microbial and vertebrate DNA. Stimulation of TLR9 induces inflammatory and invasive responses in TLR9-expressing cells. We studied here the expression of TLR9 in human MSCs and the effects of synthetic TLR9-agonists on their invasion. Constitutive expression of TLR9 was detected in human MSCs but the expression was suppressed when MSCs were induced to differentiate into osteoblasts. Using standard invasion assays and a novel organotypic culture model based on human myoma tissue, we discovered that stimulation with the TLR9 agonistic, CpG oligonucleotides increased the invasion capacity of undifferentiated MSCs. Simultaneously, an increase in MMP-13 synthesis and activity was detected in the CpG-activated MSCs. Addition of anti-MMP-13 antibody significantly diminished the CpG-induced hMSC invasion. We conclude that treatment with TLR9-ligands increases MSC invasiveness, and this process is at least partially MMP-13-mediated.

Osteoblast response to polymethyl methacrylate bioactive glass composite.

Polymethylmethacrylate (PMMA) has been used in many orthopedic and dental applications since the 1960s. Biocompatibility of newly developed surface porous fiber reinforced (SPFR) PMMA based composite has not been previously proven in cell culture environment. Analysis of rat bone marrow stromal cells grown on the different test materials showed only little difference in normalized cell activity or bone sialoprotein (BSP) production between the test materials, but the osteocalcin (OC) levels remained higher (P < 0.015-0.005) through out the test with SPFR-material when compared to tissue culture poly styrene (TCPS). The cells grown on SP-FRC material also showed highest calcium depletion from the culture medium (P < 0.026-0.001) when compared to all other test substrates. SEM images of the cultured samples confirmed that all the materials enabled cell spreading and growth on their surface, but the roughened surface remarkably enhanced this process of cell attachment, division and calcified nodule formation. This study shows that the SP-FRC composite material does not elicit harmful/toxic reactions in cell cultures more than neutral TCPS and can be considered biocompatible. The material possesses good capabilities to form new mineralized tissue onto its surface, and through that a possibility to bond directly to bone. Rough surface seems to enhance osteoblast proliferation and formation of mineralized extracellular matrix.

Bone attachment to glass-fibre-reinforced composite implant with porous surface.

A method has recently been developed for producing fibre-reinforced composites (FRC) with porous surfaces, intended for use as load-bearing orthopaedic implants. This study focuses on evaluation of the bone-bonding behaviour of FRC implants. Three types of cylindrical implants, i.e. FRC implants with a porous surface, solid polymethyl methacrylate (PMMA) implants and titanium (Ti) implants, were inserted in a transverse direction into the intercondular trabeculous bone area of distal femurs and proximal tibias of New Zealand White rabbits. Animals were sacrificed at 3, 6 and 12 weeks post operation, and push-out tests (n=5-6 per implant type per time point) were then carried out. At 12 weeks the shear force at the porous FRC-bone interface was significantly higher (283.3+/-55.3N) than the shear force at interfaces of solid PMMA/bone (14.4+/-11.0 N; p<0.001) and Ti/bone (130.6+/-22.2N; p=0.001). Histological observation revealed new bone growth into the porous surface structure of FRC implants. Solid PMMA and Ti implants were encapsulated mostly with fibrous connective tissue. Finite element analysis (FEA) revealed that porous FRC implants had mechanical properties which could be tailored to smooth the shear stress distribution at the bone-implant interface and reduce the stress-shielding effect.

Development of a multi-component fiber-reinforced composite implant for load-sharing conditions.

Fiber-reinforced composites (FRC) have the potential for use as load-bearing orthopaedic implants if the high strength and elastic modulus of FRC implant can be matched with local requirements. This study tested the in vivo performance of novel FRC implants made of unidirectional glass fibers (E-glass fibers in Bis-GMA and TEGDMA polymeric matrix). The implant surface was covered with bioactive glass granules. Control implants were made of surface-roughened titanium. Stress-shielding effects of the implants were predicted by finite element modelling (FEM). Surgical stabilization of bone metastasis in the subtrochanteric region of the femur was simulated in 12 rabbits. An oblong subtrochanteric defect of a standardized size (reducing the torsional strength of the bones approximately by 66%) was created and an intramedullary implant made of titanium or the FRC composite was inserted. The contralateral femur served as the intact control. At 12 weeks of healing, the femurs were harvested and analyzed by radiography, torsional testing, micro-CT imaging and hard tissue histology. The functional recovery was unremarkable in both groups, although the final analysis revealed two healed undisplaced peri-implant fractures in the group of FRC implants. FEM studies demonstrated differences in stress-shielding effects of the titanium and FRC implants, but the expected biological consequences did not become evident during the follow-up time of the animal study. Biomechanical testing of the retrieved femurs showed no significant differences between the groups. The torsional strength of the fixed bones had returned the level of contralateral intact femurs. Both implants showed ongrowth of intramedullary new bone. No adverse tissue reactions were observed. Based on these favorable results, a large-scale EU-project (NewBone, www.hb.se/ih/polymer/newbone) has been launched for development of orthopaedic FRC implants.

Surface porous fibre-reinforced composite bulk bone substitute.

The aim of this study was to describe and evaluate the significance of a porous surface with bioactive glass granules (S53P4) covering an artificial bulk material based on polymethylmetacrylate (PMMA) and fibre-reinforced composite (FRC) technology. Effort was focused particularly on characters of the porous surface and biomechanical properties of the material in vitro , and test in vivo the implant in reconstruction in an experimental long bone segment defect model. The defect, 10 mm in length, created in the shaft of rabbit tibia, was reconstructed by the implant and fixed by intramedullary K-wires. The implant was incorporated within 4 weeks by new bone growth from the host bone covering particularly its posterior surface and cortex/implant junctions with bridging trabecular bone. Later, at 8 weeks, new bone was found also at the cortex/implant interface and in the medullary canal of the implant. Histometric measurements revealed direct bone/implant surface contact in 34% at the interface. Bioactive glass granules in the porous surface evoked the most direct contact with bone. The implants manufactured from PMMA only served as a control group, and showed significantly lower osteoconductive properties. Biomechanical measurements in vitro of fibre-reinforced PMMA specimens revealed values for bending strength and the flexural modulus to match them to human bone. This artificial bulk bone material based on PMMA/FRC technology seems to have proposing properties to be used as a bone substitute on load-bearing conditions.

Effectiveness of multidisciplinary lifestyle intervention for hypertension: a randomised controlled trial.

Lifestyle factors like weight, alcohol consumption, salt intake and physical activity have shown to be important in treating hypertension. There have been made some randomised trials about the effects of lifestyle interventions, but the numbers of patients have been relatively small and the durations of follow-ups have been short. No controlled trials assessing the effects of lifestyle intervention in a rehabilitation setting have been reported. In this study, the effects of multidisciplinary lifestyle intervention in rehabilitation centres among middle-aged hypertensive employees were described. A total of 731 hypertensives from 45 worksites were randomised to lifestyle intervention in a rehabilitation centre or to usual care in an occupational or primary health-care centre for 12 months. Standard measurements were conducted before the intervention and 1-year later. Blood pressure (BP) levels were clearly reduced in the intervention group, while only minor changes were observed in the control group. The net changes between the two groups both for systolic and diastolic BPs were -2.1 mmHg (95% confidence intervals (CI) -4.0 to -0.1) and -1.5 mmHg (95% CI -2.6 to -0.4), respectively. The net changes were greater among men than women. The multidisciplinary lifestyle intervention in a rehabilitation centre setting produced significant reductions in BP among middle-aged employees with hypertension.

Characterization of Mycobacterium bohemicum isolated from human, veterinary, and environmental sources.

Chemotaxonomic and genetic properties were determined for 14 mycobacterial isolates identified as members of a newly described species Mycobacterium bohemicum. The isolates recovered from clinical, veterinary, and environmental sources were compared for lipid composition, biochemical test results, and sequencing of the 16S ribosomal DNA (rDNA) and the 16S-23S rDNA internal transcribed spacer (ITS) regions. The isolates had a lipid composition that was different from those of other known species. Though the isolates formed a distinct entity, some variations were detected in the features analyzed. Combined results of the phenotypic and genotypic analyses were used to group the isolates into three clusters. The major cluster (cluster A), very homogenous in all respects, comprised the M. bohemicum type strain, nine clinical and veterinary isolates, and two of the five environmental isolates. Three other environmental isolates displayed an insertion of 14 nucleotides in the ITS region; they also differed from cluster A in fatty alcohol composition and produced a positive result in the Tween 80 hydrolysis test. Among these three, two isolates were identical (cluster B), but one isolate (cluster C) had a unique high-performance liquid chromatography profile, and its gas liquid chromatography profile lacked 2-octadecanol, which was present in all other isolates analyzed. Thus, sequence variation in the 16S-23S ITS region was associated with interesting variations in lipid composition. Two of the isolates analyzed were regarded as potential inducers of human or veterinary infections. Each of the environmental isolates, all of which were unrelated to the cases presented, was cultured from the water of a different stream. Hence, natural waters are potential reservoirs of M. bohemicum.

Hospital water supply as a source of disseminated Mycobacterium fortuitum infection in a leukemia patient.

Nosocomial acquisition of Mycobacterium fortuitum led to a disseminated infection in a leukemia patient. A linkage to showerhead water was supported by molecular typing of clinical and environmental isolates. Contamination of the hospital water system with microbes that are relatively resistant to common sanitation processes poses an increased risk of infection to neutropenic patients.

Epidermolysis bullosa simplex. A new histologic subgroup.

Two patients with a new mechanobullous disease are described. The trauma-induced bullae were present at birth. The nails were deformed in both cases. Both patients were isolated cases; so far, the inheritance pattern is not known. The histologic picture was unique. The keratinocytes were dyskeratotic, enlarged, and had atypical mitoses on light microscopy of neonatal biopsy specimens. Electron microscopically the defect occurred in the tonofilaments, which formed round clumps. The blister formation took place in the lower part of the epidermis.