Diminished salivary epidermal growth factor secretion: a link between Sjögren's syndrome and autoimmune gastritis?

OBJECTIVES: Healthy human labial salivary glands produce epidermal growth factor (EGF). In Sjögren's syndrome (SS), EGF staining is diminished. SS is also associated with chronic autoimmune corpus gastritis. We therefore hypothesized that EGF secretion would be diminished in SS and that this could affect gastric target cells. METHODS: Salivary EGF secretion in SS was compared to that in healthy controls using an enzyme-linked immunosorbent assay (ELISA). EGF receptor (EGFR) immunoreactive cells in the gastric corpus of healthy human subjects were analysed using immunostaining. RESULTS: Salivary secretion of EGF was diminished in SS patients (232.4, range 52.6-618.4, vs. 756.6, range 105.3-1631.6 pg/min, p = 0.002). Proton-pump positive parietal cells were mostly EGFR immunoreactive whereas very few pepsinogen I (PGI)-positive cells were EGFR positive. CONCLUSIONS: As EGF is relatively acid resistant, salivary gland-derived EGF might participate in an exo/endocrine mode of parietal cell maintenance in the gastric corpus. Deficiency of salivary gland-derived EGF in SS patients may cause impairment of gastric parietal cells resulting in exposure of immunogenic cryptic antigens and loss of immunological self-tolerance.

Effect of S53P4 bone substitute on staphylococcal adhesion and biofilm formation on other implant materials in normal and hypoxic conditions.

To study the effect of bioactive glass bone substitute granules (S53P4) on bacterial adhesion and biofilm formation on other simultaneously used implant materials and the role of the hypoxic conditions to the adhesion. Bacterial and biofilm formation were studied on materials used both in middle ear prostheses and in fracture fixtures (titanium, polytetrafluoroethylene, polydimethylsiloxane and bioactive glass plates) in the presence or absence of S53P4 granules. The experiments were done either in normal atmosphere or in hypoxia simulating atmospheric conditions of middle ear, mastoid cavity and sinuses. We used two collection strains of Staphylococcus aureus and Staphylococcus epidermidis. In the presence of bioglass and hypoxic conditions the adhesion of the planktonic bacterial cells was decreased for most of the materials. The biofilm formation was decreased for S. epidermidis on titanium and polydimethylsiloxane in both atmospheric conditions and on bioglass plates in normoxia. For S. aureus the biofilm formation was decreased on bioglass plates and polytetrafluoroethylene in normoxia. Hypoxia produces a decrease in the biofilm formation only for S. aureus on polytetrafluoroethylene and for S. epidermidis on bioglass plates. However, in none of the cases bioactive glass increased the bacterial or biofilm adhesion. The presence of bioglass in normoxic and hypoxic conditions prevents the bacterial and biofilm adhesion on surfaces of several typical prosthesis materials in vitro. This may lead to diminishing postoperative infections, however, further in vivo studies are needed.

Effects of S53P4 bioactive glass on osteoblastic cell and biomaterial surface interaction.

To study the effect of bioactive glass bone substitute granules (S53P4) and hypoxic atmospheric conditions on human osteoblastic cell adhesion on different biomaterials. Cellular adhesion and cytoskeletal organization were studied on titanium, polytetrafluoroethylene, polydimethylsiloxane and S53P4 plates in the presence or absence of S53P4 granules. Cells used were human osteoblast-like SaOS-2 cells. The experiments were done either in normal atmospheric conditions or in hypoxia which simulates conditions prevailing in chronically infected bone or bone cavities. Vinculin-containing focal adhesions, organization of actin cytoskeleton and nuclear staining of cells on biomaterial surfaces were studied at 4.5 h, 2 and 4 days. In normoxic conditions S53P4 granules alkalinized the cell culture medium but cellular adhesion and cytoskeletal organization were usually not affected by their presence. Hypoxic conditions associated with lower pH and impaired cellular adhesion, vinculin-containing focal adhesion formation and rearrangement of the actin filaments to actin cytoskeleton. On most materials studied in hypoxic conditions, however, S53P4 granules prevented this impairment of cellular adhesion and cytoskeletal reorganization. The S53P4 granules promote the adhesion of SaOS-2 cells to various biomaterial surfaces especially in hypoxic conditions, in which S53P4 granules increase pH. The presence of S53P4 granules may protect biomaterial surface from bacterial colonization and promote osteointegration of implants used together with S53P4 granules for fixation and weight bearing.

Epithelial Cell Apoptosis in Recurrent Aphthous Ulcers.

A recurrent aphthous ulcer (RAU) is a common inflammatory ulcerative lesion affecting oral mucosa. We studied the eventual apoptosis of epithelial cells from the point of view of ulcer and inflammation. RAU lesions and healthy mucosa samples were immunostained for caspase-3 and high-mobility group box 1 (HMGB1). DNA nicks were identified using TUNEL staining. We studied the effects of tumor necrosis factor α (TNFα) and interferon γ (IFNγ) on the toll-like receptor 2 and 4 (TLR2 and TLR4) expression of human oral SCC-25 keratinocytes. We also studied the effects of self-DNA, all-thiol-HMGB1, and disulfide-HMGB1 on epithelial cells, with or without IFNγ. At the edge of RAU lesions, all epithelial cell layers were caspase-3(+), TUNEL(+), and HMGB-1(+) and had widened intercellular spaces. In contrast, healthy epithelial cells were negative for caspase-3 and TUNEL staining. HMGB1 was seen in only the basal cell layers, and the cells retained close cell-to-cell contacts. Self-DNA increased TNF-α mRNA (P = 0.02) in SCC-25 cells. Both TNFα and IFNγ (P = 0.01) increased TLR2. Upon TNFα stimulation, SCC-25 cells lost their nuclear HMGB1 staining. HMGB1 did not increase IL-8, IL-6, or TNF-α mRNA in SCC-25 cells, which was unaffected by the presence of IFNγ. We conclude that in healthy epithelium, the most superficial cells at the end of their life cycle are simply desquamated. In contrast, RAU is characterized by top-to-bottom apoptosis such that dead cells may slough off, leading to an ulcer. Because of a lack of scavenging anti-inflammatory macrophages, apoptotic cells probably undergo secondary necrosis releasing proinflammatory danger signals, which may contribute to the peripheral inflammatory halo. This is supported by self-DNA-induced TNFα synthesis. In contrast to TLR4- and TLR2-binding lipopolysaccharide used as a positive control, disulfide-HMGB1 did not stimulate proinflammatory cytokines.

Sjögren's syndome and extragonadal sex steroid formation: a clue to a better disease control?

Sjögren's syndrome (SS) is an autoimmune disease characterized by lymphoplasmacytoid focal adenitis leading to mucosal dryness, with 9:1 female dominance and peak incidence at menopause. Due to autoimmune adenitis it can be speculated that the normal epithelial cell renewal has failed, possibly as a result of local intracrine failure to process dehydroepiandrosterone (DHEA) to dihydrotestosterone (DHT). Local intracrine/-cellular DHT deficiency seems to predispose to SS if estrogens are low, in menopausal women and in men. This intracrine failure could be the initial noxious stimulus, factor X, initiating the development of SS. Abnormal release and presentation of exocrine gland-derived antigens (Ag-epitopes), in a complex with major histocompatibility complex class II (MHC II), by migratory dendritic cells (DC) activates T-cells in the regional lymph nodes. B-cells with the same specificity capture and present self-Ag to Th-cells which provide T-cell help. B-cells transform to plasma cells and start to produce autoantibodies (Ab) against these T-cell-dependent Ag. Ab against SS-A/Ro and SS-B/La ribonucleoproteins occur only in HLA-DQw2.1/DQw6 heterozygous individuals, but hY-RNA and RNA polymerase III transcripts in these Ag may in all SS patients stimulate toll-like receptors (TLR) 7 and 9 of the plasmacytoid DCs, because IFN-α and IFN-signature are produced. CD8+αEß7+cytotoxic T-cells activated via cross-presentation recirculate to attack intracrine-deficient, apoptotic epithelial cells expressing self-Ag on their surface. Exocrine glands fall into the sphere of mucosal/gut-associated lymphatic tissue. This together with immune complexes spreads the immunological memory/aggression to extra-glandular sites explaining the systemic nature of the syndrome. Secondary SS could be explained by disturbed lymphocyte recirculation. There is no conclusive evidence that SS in those few men affected is more severe than in women, suggesting that sex steroid endo-/intracrinology has its major impact on the target tissue, not on immune modulation. This article is part of a Special Issue entitled 'Essential role of DHEA'.

Histamine H4 receptor in oral lichen planus.

OBJECTIVES: Oral lichen planus (OLP) is an autoimmune disease characterized by a band-like T-cell infiltrate below the apoptotic epithelial cells and degenerated basement membrane. We tested the hypothesis that the high-affinity histamine H4 receptors (H4 Rs) are downregulated in OLP by high histamine concentrations and proinflammatory T-cell cytokines. MATERIALS AND METHODS: Immunohistochemistry and immunofluorescence staining, image analysis and quantitative real-time polymerase chain reaction of tissue samples and cytokine-stimulated cultured SCC-25 and primary human oral keratinocytes. RESULTS: H4 R immunoreactivity was weak in OLP and characterized by mast cell (MC) hyperplasia and degranulation. In contrast to controls, H4 R immunostaining and MC counts were negatively correlated in OLP (P = 0.003). H4 R agonist at nanomolar levels led to a rapid internalization of H4 Rs, whereas high histamine concentration and interferon-γ decreased HRH4 -gene transcripts. CONCLUSION: Healthy oral epithelial cells are equipped with H4 R, which displays a uniform staining pattern in a MC-independent fashion. In contrast, in OLP, increased numbers of activated MCs associate with increasing loss of epithelial H4 R. Cell culture experiments suggest a rapid H4 R stimulation-dependent receptor internalization and a slow cytokine-driven decrease in H4 R synthesis. H4 R may be involved in the maintenance of healthy oral mucosa. In OLP, this maintenance might be impaired by MC degranulation and inflammatory cytokines.

Expression of cathepsin K in periodontitis and in gingival fibroblasts.

OBJECTIVE: To study non-osteoclastic sources of cathepsin K in periodontitis. MATERIALS AND METHODS: Tissue samples were obtained from 10 otherwise healthy periodontitis pati-ents during routine periodontal flap operations and 10 systemically and periodontally healthy individuals who underwent extraction operations for retained third molars. Methods used were immunohistochemistry, image analysis, immunofluorescence double-staining, gingival fibroblast culture, tumour necrosis factor-α (TNF-α) stimulation and Western blotting. RESULTS: Macrophage-like cells, fibroblast-like cells, vascular endothelial cells and gingival epithelial cells were more intensively stained for cathepsin K and also more frequent in periodontitis than in controls (665 ± 104 vs 258 ± 40 cells mm(-2) , P < 0.01). Some cathepsin K(+) cells in periodontal tissues were CD68(+) , but some were CD68(-) and probably fibroblasts. Indeed, in gingival fibroblast culture, resting fibroblasts released cathepsin K, more 43 kD procathepsin K than 29 kD active cathepsin K. TNF-α increased the release of the activated cathepsin K 4- to 5-fold. CONCLUSIONS: Results suggest that GCF-cathepsin K is not only osteoclast-derived, but in periodontitis, also other cells contribute to it. GCF-cathepsin K, perhaps together with intracellular, lysosomal collagenolytically active cathepsin K in fibroblasts, macrophages and gingival epithelial cells, can contribute to the loss of attachment and destruction of the periodontal ligament.

Increased beta 2 defensin in recurrent aphthous ulcer.

OBJECTIVES: It was hypothesized that beta 2 defensin (BD-2) is increased in RAU lesions compared with healthy controls to promote anti-microbial host defence. METHODS: RAU and control mucosa samples were subjected to quantitative real-time PCR and immunostained for BD-2, CD68, mast cell tryptase and 4-hydroxynonenal (4HNE). The effect of tumour necrosis factor-α (TNF-α) ± interleukin-17C (IL-17C), without and with vitamin K3, was studied on BD-2 expression in epithelial SCC-25 cells. RESULTS: Although BD-2 mRNA did not differ between healthy and RAU mucosa, BD-2 stained strongly in acute-phase RAU epithelium (P = 0.001). In controls, subepithelial BD-2(+) cells were mast cells and macrophages, whereas in RAU, most infiltrating leucocytes were BD-2(+) (P = 0.004). In cell culture, BD-2 was increased 124-fold by TNF-α (P < 0.0001) and 208-fold synergistically together with IL-17C (P < 0.0001). 4HNE staining of RAU epithelium was not significantly increased, and vitamin K3-induced reactive oxygen species (ROS) did not affect BD-2. CONCLUSIONS: Anti-microbial BD-2 was not affected by oxidative stress but was highly increased in the epithelial and immigrant cells in the acute-phase RAU lesions, probably in part synergistically by TNF-α and epithelial IL-17C, which are known to be induced by activation of danger-signal receptors by pathogen- and/or damage-associated molecular patterns.

Osteoarthritic cartilage lesions in the bovine patellar groove: a macroscopic, histological and immunohistological analysis.

A high percentage of osteoarthritis (OA)-like patellar groove lesions in the stifle joint in calcium-deficient bulls has been recently reported. The prevalence of these lesions in bulls deficient in or supplemented with calcium was compared to findings in culled and healthy bulls to determine whether they represent normal anatomical variations, developmental anomalies or OA. It was hypothesized that the patellar groove lesions may represent OA. Distal cartilage samples from 160 femurs were analysed using a macroscopic Société Française d'Arthroscopie (SFA) OA grading system. Samples representing different SFA grades were subjected to Osteoarthritis Research Society International (OARSI) histological and high-mobility group box 1 (HMGB1) immunohistological OA grading. For a qualitative analysis three OA samples were immunostained for interleukin (IL)-1ß, matrix metalloproteinase (MMP)-13 and collagenase-produced COL2-3/4M neoepitopes. Patellar groove lesions were found in 48% of the femurs and were highest in calcium-deficient animals (71%, P<0.001). All three different grading systems disclosed OA in culled bulls, but no focal areas of cartilage necrosis. OARSI and HMGB1 grades were fairly concordant (Spearman's ρ=0.95, P<0.001; Cohen's κ=0.23, P<0.005), both with a slight disparity with the SFA grade (ρ=0.80 and 0.87, P<0.01; κ=0.36 and 0.46, P<0.001). IL-1ß, MMP-13 and COL2-3/4M staining patterns were compatible with OA. The study showed that patellar groove lesions are common in bulls. In all SFA, OARSI and HMGB1 graded samples the lesions clearly demonstrated OA and showed OA-typical pathophysiology. Arthroscopic SFA grading showed similar changes in calcium-deficient and calcium-supplemented bulls, but in the absence of a time course study and histological data the primary nature of these lesions could not be established with certainty.

BMP-2 induces ATF4 phosphorylation in chondrocytes through a COX-2/PGE2 dependent signaling pathway.

OBJECTIVE: Bone morphogenic protein (BMP)-2 is approved for fracture non-union and spine fusion. We aimed to further dissect its downstream signaling events in chondrocytes with the ultimate goal to develop novel therapeutics that can mimic BMP-2 effect but have less complications. METHODS: BMP-2 effect on cyclooxygenase (COX)-2 expression was examined using Real time quantitative PCR (RT-PCR) and Western blot analysis. Genetic approach was used to identify the signaling pathway mediating the BMP-2 effect. Similarly, the pathway transducing the PGE2 effect on ATF4 was investigated. Immunoprecipitation (IP) was performed to assess the complex formation after PGE2 binding. RESULTS: BMP-2 increased COX-2 expression in primary mouse costosternal chondrocytes (PMCSC). The results from the C9 Tet-off system demonstrated that endogenous BMP-2 also upregulated COX-2 expression. Genetic approaches using PMCSC from ALK2(fx/fx), ALK3(fx/fx), ALK6(-/-), and Smad1(fx/fx) mice established that BMP-2 regulated COX-2 through activation of ALK3-Smad1 signaling. PGE-2 EIA showed that BMP-2 increased PGE2 production in PMCSC. ATF4 is a transcription factor that regulates bone formation. While PGE2 did not have significant effect on ATF4 expression, it induced ATF4 phosphorylation. In addition to stimulating COX-2 expression, BMP-2 also induced phosphorylation of ATF4. Using COX-2 deficient chondrocytes, we demonstrated that the BMP-2 effect on ATF4 was COX-2-dependent. Tibial fracture samples from COX-2(-/-) mice showed reduced phospho-ATF4 immunoreactivity compared to wild type (WT) ones. PGE2 mediated ATF4 phosphorylation involved signaling primarily through the EP2 and EP4 receptors and PGE2 induced an EP4-ERK1/2-RSK2 complex formation. CONCLUSIONS: BMP-2 regulates COX-2 expression through ALK3-Smad1 signaling, and PGE2 induces ATF4 phosphorylation via EP4-ERK1/2-RSK2 axis.

Novel biological strategies for treatment of wear particle-induced periprosthetic osteolysis of orthopaedic implants for joint replacement.

Wear particles and by-products from joint replacements and other orthopaedic implants may result in a local chronic inflammatory and foreign body reaction. This may lead to persistent synovitis resulting in joint pain and swelling, periprosthetic osteolysis, implant loosening and pathologic fracture. Strategies to modulate the adverse effects of wear debris may improve the function and longevity of joint replacements and other orthopaedic implants, potentially delaying or avoiding complex revision surgical procedures. Three novel biological strategies to mitigate the chronic inflammatory reaction to orthopaedic wear particles are reported. These include (i) interference with systemic macrophage trafficking to the local implant site, (ii) modulation of macrophages from an M1 (pro-inflammatory) to an M2 (anti-inflammatory, pro-tissue healing) phenotype in the periprosthetic tissues, and (iii) local inhibition of the transcription factor nuclear factor kappa B (NF-κB) by delivery of an NF-κB decoy oligodeoxynucleotide, thereby interfering with the production of pro-inflammatory mediators. These three approaches have been shown to be viable strategies for mitigating the undesirable effects of wear particles in preclinical studies. Targeted local delivery of specific biologics may potentially extend the lifetime of orthopaedic implants.

No evidence of long-term benefits of arthroscopicacromioplasty in the treatment of shoulder impingement syndrome: Five-year results of a randomised controlled trial.

OBJECTIVES: To report the five-year results of a randomised controlled trial examining the effectiveness of arthroscopic acromioplasty in the treatment of stage II shoulder impingement syndrome. METHODS: A total of 140 patients were randomly divided into two groups: 1) supervised exercise programme (n = 70, exercise group); and 2) arthroscopic acromioplasty followed by a similar exercise programme (n = 70, combined treatment group). RESULTS: The main outcome measure was self-reported pain as measured on a visual analogue scale. At the five-year assessment a total of 109 patients were examined (52 in the exercise group and 57 in the combined treatment group). There was a significant decrease in mean self-reported pain on the VAS between baseline and the five-year follow-up in both the exercise group (from 6.5 (1 to 10) to 2.2 (0 to 8); p < 0.001) and the combined treatment group (from 6.4 (2 to 10) to 1.9 (0 to 8); p < 0.001). The same trend was seen in the secondary outcome measures (disability, working ability, pain at night, Shoulder Disability Questionnaire and reported painful days). An intention-to-treat analysis showed statistically significant improvements in both groups at five years compared with baseline. Further, improvement continued between the two- and five-year timepoints. No statistically significant differences were found in the patient-centred primary and secondary parameters between the two treatment groups. CONCLUSIONS: Differences in the patient-centred primary and secondary parameters between the two treatment groups were not statistically significant, suggesting that acromioplasty is not cost-effective. Structured exercise treatment seems to be the treatment of choice for shoulder impingement syndrome.

Osteolysis around total knee arthroplasty: a review of pathogenetic mechanisms.

Aseptic loosening and other wear-related complications are some of the most frequent late reasons for revision of total knee arthroplasty (TKA). Periprosthetic osteolysis (PPOL) pre-dates aseptic loosening in many cases, indicating the clinical significance of this pathogenic mechanism. A variety of implant-, surgery- and host-related factors have been delineated to explain the development of PPOL. These factors influence the development of PPOL because of changes in mechanical stresses within the vicinity of the prosthetic device, excessive wear of the polyethylene liner, and joint fluid pressure and flow acting on the peri-implant bone. The process of aseptic loosening is initially governed by factors such as implant/limb alignment, device fixation quality and muscle coordination/strength. Later, large numbers of wear particles detached from TKA trigger and perpetuate particle disease, as highlighted by progressive growth of inflammatory/granulomatous tissue around the joint cavity. An increased accumulation of osteoclasts at the bone-implant interface, impairment of osteoblast function, mechanical stresses and increased production of joint fluid contribute to bone resorption and subsequent loosening of the implant. In addition, hypersensitivity and adverse reactions to metal debris may contribute to aseptic TKA failure, but should be determined more precisely. Patient activity level appears to be the most important factor when the long-term development of PPOL is considered. Surgical technique, implant design and material factors are the most important preventative factors, because they influence both the generation of wear debris and excessive mechanical stresses. New generations of bearing surfaces and designs for TKA should carefully address these important issues in extensive preclinical studies. Currently, there is little evidence that PPOL can be prevented by pharmacological intervention.

Urban legends series: Sjögren's syndrome.

Sjögren's syndrome (SjS) is one of the most common autoimmune rheumatic diseases, clinically characterized by xerostomia and keratoconjunctivitis sicca. We investigated the following controversial topics: (i) Do we have reliable ways of assessing saliva production? (ii) How important are the quantity and quality of saliva? (iii) Are only anti-SSA/Ro and anti-SSB/La relevant for the diagnosis of SjS? (iv) Are the American-European Consensus criteria (AECC) the best way to diagnose SjS? Results from literature searches suggested the following: (i) Despite the fact that numerous tests are available to assess salivation rates, direct comparisons among them are scarce with little evidence to suggest one best test. (ii) Recent developments highlight the importance of investigating the composition of saliva. However, more research is needed to standardize the methods of analysis and collection and refine the quality of the accumulating data. (iii) In addition to anti-Ro/La autoantibodies, anti α-fodrin IgA and anti-MR3 autoantibodies seem to be promising diagnostic markers of SjS, but more studies are warranted to test their sensitivity and specificity. (iv) AECC are classification, not diagnostic criteria. Moreover, recent innovations have not been incorporated into these criteria. Consequently, treatment directed to patients diagnosed using the AECC might exclude a significant proportion of patients with SjS.

Bovine chronic osteoarthritis causes minimal change in synovial fluid.

Chronic osteoarthritis (OA) is a degenerative disease of the articular cartilage. DNA-binding high mobility group protein B1 (HMGB1) is released on cellular death/activation and acts as an endogenous danger signal and a proinflammatory cytokine. Matrix metalloproteinase (MMP)-2 and in MMP-9 are induced to mediate proteolytic degradation/remodelling of joint tissues. Collagen degradation in the bone and synovium leads to release of type I collagen-derived cross-linked carboxy-terminal telopeptide (ICTP). These molecules have been linked to the pathogenesis of OA and could have potential as synovial fluid (SF) biomarkers in OA. Cartilage and SF were obtained from 27 dairy bulls (30-61 months old) and control cartilage from six young healthy dairy bulls. OA lesions were evaluated grossly (five grades), histologically (seven Osteoarthritis Research Society International [ORSI] grades) and immunohistochemically (four HMGB1 grades). The OARSI lesion score was calculated as the product of the OARSI grade and the OARSI score (the total area of the lesions). SF concentrations of HMGB1, MMP-2 and -9 and ICTP were measured by enzyme-linked immunosorbent assay, gelatin zymography and radioimmunoassay, respectively. Seventy-two percent (39/54) of stifle joints and 85% (23/27) of the dairy bulls had at least one gross OA lesion and 94% of the lesions were localized to the distal end of the femur, with the patellar groove and the lateral trochlear ridge being predilection sites. Gross and histological grades correlated with the HMGB1 grade, but SF total cell count, percent neutrophils or the measured biomarkers did not correlate with the tissue lesions, with the exception of ICTP concentration, which correlated with the total joint score. The switch of HMGB1 from DNA-binding nuclear protein to an extracellular alarmin/cytokine correlates with the gross and histological grades of OA tissue lesions. However, the activity and extent of the tissue lesions did not correlate with other SF biomarkers, perhaps because the histological grades represent outcome measures, while SF reflects process parameters. The only exception was ICTP concentration, which reflects enhanced destruction/remodelling.

Spectroscopy in the analysis of bacterial and eukaryotic cell footprints on implant surfaces.

We tested the suitability of two spectroscopic methods, x-ray photoelectron spectroscopy (XPS) and time of flight secondary ion mass spectrometry (ToF-SIMS), in the recognition of bacterial and eukaryotic cell footprints on implant surfaces. Human mesenchymal stem cells (MSCs) and Staphylococcus aureus were cultured on sample surfaces and detached using trypsin. Scanning electron microscopy confirmed that the processed surfaces did not contain any human or microbial cells. The footprints were then analysed using XPS and ToF-SIMS. XPS results showed no significant differences between the footprints, but principal component analysis of the ToF-SIMS data enabled clear separation of MSC-footprints from the S. aureus and co-culture footprints (p < 0.03). ToF-SIMS also demonstrated 'race for the surface' between proteins, which suggest surface charge (zeta-potential) dependent protein adsorption. ToF-SIMS differentiated eukaryotic and bacterial footprints and has potential for post-hoc detection of implant-related infections based on the typical ToF-SIMS spectra.

RANKL in the osteolysis of AES total ankle replacement implants.

Peri-implant tissue reactions in failed total ankle replacement (TAR) are characterized by early developing peri-implant osteolysis. The hypothesis of the study was that this reaction is mediated by receptor activator of nuclear factor kappa B ligand (RANKL). Samples of peri-prosthetic tissues from failed TAR implants were stained for macrophages, RANKL, its receptor RANK and osteoprotegerin (OPG), and compared to control samples. The failed TAR implants were surrounded by implant capsule, synovial lining-like interface membrane or necrotic tissues. Infiltrating scavenger receptor I positive CD163(+) macrophages were frequent, in particular around necrotic soft tissues or bone sequestrate, and possibly in part formed due to ischemia and mechanical factors. In contrast, implant-derived wear debris was scanty. Still many RANK(+) macrophages were often seen in close contact with RANKL(+) mesenchymal cells, whereas OPG was mostly located at a distance in vascular endothelial cells. Foreign body giant cells were frequent. RANKL seems to stimulate locally accumulated CD163(+) RANK-expressing cells to fusion, which leads to the local formation of multinuclear foreign body giant cells (and probably of osteoclasts). Therefore, peri-implant osteolysis in early TAR implant failure seems to be caused by the RANKL-driven chronic foreign body inflammation directed against, not implant-derived particles, but against necrotic autologous tissues.

Brief report: first identification of H4 histamine receptor in healthy salivary glands and in focal sialadenitis in Sjögren's syndrome.

OBJECTIVE: The conventional H(1) and H(2) histamine receptors have >10,000-fold lower avidity for histamine than H(4) histamine receptor, which has been implicated in autoimmune diseases. This study was undertaken to compare H(4) histamine receptor levels in the salivary glands (SGs) of healthy controls with those in the SGs of patients with primary Sjögren's syndrome (SS). METHODS: H(4) histamine receptor messenger RNA (mRNA) was analyzed using real-time quantitative polymerase chain reaction, and the receptor protein was examined using immunostaining. Effects of the H(4) histamine receptor agonist ST-1006 on cytokine synthesis by human SG (HSG) cells were analyzed using xMAP technology and enzyme-linked immunosorbent assay. RESULTS: Healthy SGs contained H(4) histamine receptor mRNA. The receptor protein was localized to the acinar and ductal epithelial cells. H(4) histamine receptor agonist stimulated HSG cells to produce the cytokines interleukin-8 and vascular endothelial growth factor. SS patients had low H(4) histamine receptor levels. CONCLUSION: H(1) and H(2) histamine receptor antagonists are not effective in the treatment of autoimmune diseases. However, such antagonists do not affect the newly discovered H(4) histamine receptor. Dendritic cells and lymphocytes are nonprofessional histamine-producing cells, which produce histamine at 100-1,000-fold lower rates than mast cells do. Saliva contains only 0.31-12.4 ng/ml histamine, which is too low to stimulate H(1) or H(2) histamine receptor, but stimulates H(4) histamine receptor half maximally. Our findings show that H(4) histamine receptor is strongly expressed in tubuloacinar SG cells, which emphasizes the role of these cells in the pathogenesis of SS.

Effect of oral clodronate on structural damage and bone turnover in rheumatoid arthritis.

OBJECTIVES: To study the effect of oral clodronate on structural damage and bone metabolism in rheumatoid arthritis (RA). METHODS: In this 2-year proof-of-concept study, sixty patients with at least moderately active RA were randomised to receive anti-rheumatic therapy alone or together with oral clodronate 1600 mg daily. Radiographs of hands and feet and serum samples for bone biomarkers were studied at baseline and at 24 months. RESULTS: At 24 months, progression of radiographic joint damage was similar in the 2 groups. Clodronate suppressed carboxyterminal cross-linked peptide of type I collagen (p=0.03) and aminoterminal propeptide of type I procollagen (p=0.01). Eight patients (27%) withdrew from clodronate therapy due to adverse drug reactions. CONCLUSIONS: Oral clodronate did not retard the focal bone damage in RA despite its beneficial effect on overall bone metabolism, as judged by the decrease in the reference bone biomarkers.