Circulating miRNAs in hand osteoarthritis.

OBJECTIVE: Hand osteoarthritis (OA) is a frequent musculoskeletal disorder with an increasing prevalence during ageing. This study aimed to evaluate circulating microRNAs (miRNAs) in the plasma of patients with hand OA compared with age- and sex-matched healthy control subjects. METHODS: In total, 238 participants (96 with erosive and 73 with non-erosive hand OA patients and 69 healthy control subjects) were included in this study. All patients underwent clinical examinations, including self-reported measures (AUSCAN and Algofunctional index). Radiographs of both hands were scored with the Kallman scale. The profile of miRNAs in plasma was screened using TaqMan™ Low-Density Array, and candidate miRNAs were validated on two quantitative real-time PCR (qRT-PCR) systems (QuantStudio and SmartChip). RESULTS: Of all the 754 miRNAs, 40 miRNAs were different between hand OA patients and healthy control subjects in the screening cohort. Following the two-phase validation process, three miRNAs (miR-23a-3p, miR-146a-5p, and miR-652-3p) were increased in patients with hand OA compared with healthy control subjects and were associated with the AUSCAN sum score and AUSCAN pain. Furthermore, an inverse correlation of miR-222-3p with the Kallman radiographic score was found. The expression of miRNAs did not differ between erosive and non-erosive hand OA. CONCLUSION: The profile of circulating miRNAs could unveil candidate biomarkers associated with hand OA symptoms. Longitudinal studies are required to determine the role of miRNAs in hand OA.

Elevated Tenascin-C Serum Levels in Patients With Axial Spondyloarthritis.

This study aimed to examine serum tenascin C (TNC) in different subsets of axial spondyloarthritis (axSpA) patients. Sixty-one patients fulfilling the Assessment of SpondyloArthritis international Society classification criteria for axSpA and 20 healthy subjects (HS) were included in study. Based on imaging, patients were classified as non-radiographic (n=16) and radiographic (n=45) axSpA. TNC serum levels were determined by ELISA. Disease-related factors including the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) and C-reactive protein (CRP) levels, were determined. TNC levels were elevated in axSpA patients [535.3 (457.7-677.2) ng/ml] compared to HS [432.1 (329.1-565.9) ng/ml, p=0.007]. Dividing axSpA into radiographic and non-radiographic subsets, the difference in TNC was observed between the radiographic subset and HS [535.3 (434.5-677.2) vs. 432.1 (329.1-565.9) ng/ml, p=0.022]. TNC levels did not correlate with disease activity measures (serum CRP or BASDAI). Nevertheless, the weak correlation of TNC levels with different disease stages (r=0.25, p=0.025) was found, with the highest levels in patients with syndesmophytes. TNC levels are elevated across various subsets of axSpA, and although not related to systemic disease activity, TNC levels might reflect chronic structural spinal changes in axSpA patients. However, its specific role in bone metabolism should be elucidated in further studies.

Is serum TWEAK a useful biomarker of neuropsychiatric systemic lupus erythematosus?

The aim of this study was to determine the role of the tumor necrosis factor like weak inducer of apoptosis (TWEAK) as a serum biomarker of neuropsychiatric involvement in systemic lupus erythematosus (NPSLE). Levels of TWEAK levels were measured in sera of 92 patients with systemic lupus erythematosus (SLE), including 28 patients with neuropsychiatric lupus, and in 59 healthy controls using ELISA. All SLE patients underwent rheumatological, neurological and psychiatric assessment. We found no significant differences in TWEAK levels, between SLE patients and the healthy controls (p=0.2411). Similarly, no difference was observed between subgroup of NPSLE and healthy controls (p=0.7658). The mean SLE disease activity (SLEDAI) was 13.25. No correlations between TWEAK levels with disease activity (SLEDAI, r=0.2113, p=0.2805) or the most common NPSLE manifestations such as headache (r=0.2079), seizures (r=0.1101), cerebrovascular disease (r= 0.2347), cognitive dysfunction (r=0.1597) and anxiety (r=0.1397) were observed. Our data do not support the use of serum TWEAK as a discriminating biomarker for NPSLE. The role of the TWEAK in NPSLE remains to be investigated.

Decrease in serum interleukin-21 levels is associated with disease activity improvement in patients with recent-onset rheumatoid arthritis.

Interleukin-21 (IL-21) plays an important role in the pathogenesis of rheumatoid arthritis (RA). The aim of our study was to assess serum levels of IL-21 in patients with recent-onset RA in relation to disease activity and response to treatment. We analyzed serum levels of IL-21 in 51 RA patients, both before and 12 weeks after the initiation of treatment and in 36 healthy individuals. Disease activity was assessed at baseline and at weeks 12 and 24 using the Disease Activity Score for 28 joints, serum levels of C-reactive protein, and the total swollen joint count. We found that IL-21 levels were not increased in patients with recent-onset RA compared with healthy controls, but they had significantly decreased from baseline to week 12 during treatment. Baseline levels of IL-21 significantly correlated with measures of disease activity (p<0.02 for all). Although IL-21 levels did not predict achievement of remission, decrease in IL-21 levels correlated with improvement in disease activity after 12 weeks (p<0.02) and also after 24 weeks (p<0.04) of treatment. Our data suggest that circulating IL-21 levels may serve as a biomarker of disease activity and better outcome in early phase of RA.

Novel adipokine fibroblast growth factor 21 is increased in rheumatoid arthritis.

Fibroblast growth factor-21 (FGF-21) has been recently characterized as a new adipokine. The aim of this study was to assess FGF-21 levels in patients with rheumatoid arthritis (RA) and osteoarthritis (OA) and to study the relationship between FGF-21, disease activity and metabolic status. The levels of FGF-21 in serum and synovial fluid samples from 38 patients with RA and 42 control individuals with OA were determined by ELISA. Patients were assessed for disease activity using the disease activity score (DAS28), a serum glucose and lipid profile. Age, sex and BMI-adjusted FGF-21 levels in the serum (p=0.024) and synovial fluid (p=0.010) samples were significantly higher in patients with RA when compared with OA. The levels of FGF-21 in the serum significantly correlated with the levels in the synovial fluid. Serum and synovial fluid FGF-21 levels adjusted for confounders correlated positively with C-reactive protein. The levels of FGF-21 were positively correlated with BMI in patients with RA; however, the levels were not associated with disease activity or lipid profiles. Furthermore, serum FGF-21 levels were significantly higher in seropositive compared with seronegative RA patients. This work shows that patients with seropositive RA have increased levels of FGF-21. The results suggest that FGF-21 is related to BMI but not disease activity or lipid profiles in patients with RA.

Low circulating Dickkopf-1 and its link with severity of spinal involvement in diffuse idiopathic skeletal hyperostosis.

OBJECTIVE: Dickkopf-1 (DKK-1) is an inhibitor of osteoblastogenesis, and its lower levels are linked to new bone formation. The aim of this study was therefore to explore serum levels of DKK-1 and to evaluate DKK-1's association with the severity of spinal involvement in diffuse idiopathic skeletal hyperostosis (DISH). METHODS: Serum levels of total and functional DKK-1 and C-reactive protein (CRP) were measured in 37 patients with DISH and 22 healthy age and sex-matched controls. Plain radiographs of the cervical and thoracic spine were performed, and the diagnosis of DISH was defined using the Resnick criteria. Patients were divided into three groups based on spinal involvement. Bone mineral density (BMD) and bone turnover markers were evaluated in patients with DISH. RESULTS: The levels of total serum DKK-1 were significantly lower in patients with DISH than in healthy controls (p<0.0001). Importantly, low serum levels of DKK-1 were associated with more severe spinal involvement in DISH, independent of age, sex, disease duration, CRP, bone turnover markers or BMD. However, these findings were less significant for functional DKK-1. CONCLUSION: These observations indicate that DKK-1 may play a significant role in bone formation during DISH.

Surface modification of cyclic olefin copolymers for osteochondral defect repair can increase pro-destructive potential of human chondrocytes in vitro.

Materials on the basis of cycloolefin copolymers (COC) are suitable for subchondral defect repairs. The objective of this study was to evaluate the influence of surface modification of COC and COC/LLDPE blends on the viability and gene expression of chondrocytes. Human chondrocytes were incubated on the surface of the studied materials. Half of the materials were plasmatically modified with a subsequent type II collagen application. The gene expression of matrix metalloproteinases (MMP-1,-3,-13), pro-inflammatory cytokines (IL-1, TNF-alpha) and apoptotic molecules (BAX, Bcl-2) was evaluated using quantitative Taq-Man PCR after 48 h incubation. Chondrocyte viability was evaluated by the MTT test after 2, 4 and 8 days of incubation. The synthesis of MMPs was measured by ELISA assay in cell culture medium after 48 h of incubation. Chondrocytes incubated on plasmatically modified in contrast to unmodified materials demonstrated significantly increased gene expression of IL-1 (p<0.05), MMP-1 and MMP-3 (p<0.05 for both comparisons) as well as MMP-13 (p<0.001). Increased gene expression was confirmed by significantly increased production of active forms of particular MMPs into the cell culture medium. Unlike surface unmodified polymers, the modified materials showed time-dependent reduction of chondrocyte viability. The gene expression of TNF-alpha and apoptotic molecules by chondrocytes was not significantly changed by different materials. Cycloolefin copolymers and their blends may represent suitable materials for tissue engineering, however, their surface modification followed by collagen type II application may, at least under in vitro conditions, reduce the viability of chondrocytes and induce their pro-destructive behavior. The potential benefit or disadvantage of surface modifications of materials for osteochondral defect repairs needs to be further elucidated.

Serum hyaluronic acid as a potential marker with a predictive value for further radiographic progression of hand osteoarthritis.

OBJECTIVE: To compare serum levels of hyaluronic acid (HA) between patients with erosive and non-erosive hand osteoarthritis (HOA), and investigate its association with morphological changes and radiographic progression over 2 years. METHODS: Fifty-five women with erosive and 33 women with non-erosive HOA were included in this study. All underwent clinical examination, which included assessment of pain, swelling, deformity and deviation of small hand joints and completed health assessment questionnaires. Serum levels of HA were measured by ELISA. Three-phase bone scintigraphy was performed at baseline. Radiographs of both hands were performed at baseline and after 2 years and scored according Kallman grading scale. RESULTS: Serum levels of HA were significantly higher in patients with erosive than with non-erosive HOA (P<0.01). It correlated significantly with the number of hand joints with deviations and deformities. HA adjusted for age and disease duration significantly correlated with radiographs at baseline and after 2 years in all patients with HOA (r=0.560 and r=0.542, P<0.01 for both correlations). Although there was an association between HA and radiographic score in erosive disease, after adjustment for confounders it remained no longer significant. HA adjusted for confounders correlated significantly with the late phase in all patients with HOA (r=0.412, P<0.01) and in patients with erosive disease (r=0.320, P<0.05). CONCLUSION: HA is increased in patients with erosive HOA and could be proposed as a surrogate marker with a predictive value for further radiographic progression of HOA in general. Further investigation is necessary to confirm these results.

Collagen degradation products and proinflammatory cytokines in systemic and localized scleroderma.

The aim of this study was to assess the degradation of collagen type I and proinflammatory cytokines in systemic and localized scleroderma compared with psoriasis and healthy controls. Total 99 individuals were examined - 24 with SSc, 22 with LSc, 39 patients with PsV and 14 healthy controls. U-PD and U-DPD were measured using a sensitive isocratic HPLC method. Serum levels of IL-6 and soluble IL-2R were assayed using commercial ELISA kits. In the SSc group U-PD and U-DPD levels (nmol/mmol creatinine) were increased compared with controls (P = 0.001) and with PsV (P = 0.006). IL-6 levels were increased compared with controls (P = 0.004) and with PsV (P = 0.002). IL-2R concentrations were insignificantly increased in comparison with controls and were lower than in PsV, but the difference was not significant. In the LSc group excretion of U-PD and U-DPD did not differ from controls, but was insignificantly decreased compared with PsV. IL-6 levels were increased compared with controls (P = 0.001) and also with PsV (P = 0.03). IL-2R concentrations were significantly increased in comparison with controls only (P = 0.03). In patients with SSc our data have shown the most intensive collagen degradation and simultaneously an active inflammation, as documented by IL-6, which reflects the pathological processes in the skin and visceral organs compared with PsV patients and healthy individuals. In the LSc group collagen degradation was similar to that in control groups, but a certain inflammatory activity was observed.

The influence of implant surface properties on cell adhesion and proliferation.

Interactions of the foreign material of implant and the living tissue on the cell level can cause prolonged healing or, worse, loss of the implant. The cell response to the presence of some implant materials was studied under in vitro conditions. The influence of physicochemical surface parameters on the response of the cells in the immediate vicinity of implants, namely on adhesion, proliferation and synthetic activity of fibroblasts, and on the blood coagulation were compared. The direct contact of tested materials (titanium and Ti6Al4V alloy with various surface treatments, Cr Co Mo alloy, hydroxyapatite-coated titanium, zirconium oxide ceramics, polyethylene and carbon composite) on cell spreading was monitored and the presence of TNF-alpha and IL-8 was evaluated in the cultivation medium. The formation of blood clots was investigated on samples immersed in a well with freshly drawn whole rabbit blood using a scanning electron microscope. The surface free energy was estimated using the measurement of static contact angle. Both the advancing and receding contact angles were measured by the dynamic Wilhemy plate method. Two main groups with extremes in cell viability were established. In the first group the increased polar component of surface free energy, the highest cell density, the lowest inflammatory cytokine production, but no fibres in the clotting blood were found. On the contrary, the second group of materials with a very low polar component of the surface free energy showed distinctly higher expression of inflammatory mediators, low cell proliferation, but faster formation of fibres in the blood coagulum.

Clinical correlations of potential activity markers in systemic sclerosis.

The objectives of the project were the following: (1) to establish a group of patients with a confirmed diagnosis of systemic sclerosis (Ssc), (2) to perform a detailed entrance examination of each patient, (3) to determine concentrations of potential activity markers, and (4) to make a comprehensive examination of each patient 1 year after inclusion into the study. A total of 49 patients were examined, 36 with a limited form of SSc, 9 with diffuse SSc, and 4 with other forms of SSc. We determined plasma or serum levels of the N-terminal propeptide of procollagen type III (NPIIIP), interleukin-6 (IL-6), soluble receptor for interleukin-2 (sIL-2r), soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular adhesion molecule-1 (sVCAM-1), von Willebrand factor antigen (vWFAg), and big endothelin-1 (BET-1) using commercial kits, and urinary excretion of pyridinoline (PYR) and deoxypyridinoline (D-PYR) using high-performance liquid chromatography. Correlations of these markers with selected clinical data were calculated. The mean levels of all potential activity markers were increased compared with normal values, but differences were not significant. The levels of NPIIIP, D-PYR, and IL-6 were normal. The measured values after 1 year did not differ from the entry values. At entry, NPIIIP concentrations correlated with the finger-to-palm distance, and D-PYR corresponded with findings on a simplified health assessment questionnaire (FQ). IL-6 levels correlated with the leukocyte count, sIL-2r with the FQ, and ET-1 with the diffuse lung capacity for carbon monoxide. In general, we found only a few clinical correlates of potential activity markers. Our data confirmed the correlations of collagen metabolism markers with skin involvement and FQ, as was reported previously. Larger studies in this field are needed.

Biological properties of the intervertebral cages made of titanium containing a carbon-carbon composite covered with different polymers.

Intervertebral cages are used in orthopaedics for stabilization of injured lumbar parts of vertebral columns. Our study provides preliminary results of tests of the biological properties of titanium cages with a variously modified carbon/carbon composite (C/C) core. This core was produced from a C/C composite modified by hydrogel materials based on poly(2-hydroxyethyl methacrylate) (HEMA) enriched with 1% collagen or 35% methylmethacrylate or 30% terc-butylmethacrylamide. We evaluated the adhesion of the cells to the tested material coating using an in vitro study of the metabolic activity and cytokine production of the cells (TNF-alpha, IL-8). We studied the biocompatibility of intervertebral cages coated with different copolymers under in vivo condition and in an implantation experiment in the porcine femurs. Both in vitro and in vivo results revealed favourable biotolerance of the use system. Modification of the composite HEMA with the use of collagen seems to have a more positive effect on the new bone tissue formed around the implanted devices than HEMA copolymerized with methylmethacrylate or terc-butylmethacrylamide.

[Changes in the articular compartment in advanced osteoarthritis]. / Zmeny v kloubním kompartmentu u pokrocilé osteoartrózy.

PURPOSE OF THE STUDY: Both synovial and bone forms of osteoarthritis (OA) are characterized by inflammatory processes in the articular compartment. Increasing evidence suggests that changes in bone tissue are important for the deterioration or loss of joint function. Therefore it is reasonable to shift emphasis from research on cartilage to that on other articular tissues, particularly on subchondral bone. The aim of the study was to demonstrate the involvement of several cytokines in OA development and, on the basis of changes of joint markers, to assess the extent of inflammatory process. MATERIAL: A total of 60 patients with an osseous form of osteoarthritis of the knee joint (28 patients) or the hip joining (32 patients) underwent total knee or hip arthroplasty. The mean age of our patients was 66.7 +/- 10.4 years. Preoperative clinical and radiographic examinations were carried out as well as routine laboratory tests on blood and urine. Samples of urine, blood serum (BS) and synovial fluid (SF), extracts from cartilage (CA) and synovial membrane (SM) and granulation bone tissue were analysed for markers indicating the presence of inflammatory processes in joints. METHOD: The following markers of inflammatory activity in the bone compartment were investigated: pyridinoline (PYR), deoxypyridinoline (D-PYR), bone alkaline phosphatase (BAP) and chondrex (CHON). The levels of cytokines IL-1 alpha, IL-8, IL-10 and TNF-alpha were assayed by immunoanalysis (ELISA and IMMULITE system) in BS, CA, SM, GT and SF. The tissue samples were obtained during arthroplasty. RESULTS: In the patients with osteoarthritis, the urinary levels of PYR and D-PYR were higher than control values (70.33 +/- 34.93 vs (41.6 +/- 10.6 nmol/mmol creatinine). No significant differences were found between pre- and post-operative levels. Similarly, the serum levels of BAP and CHON compared with control values were higher (27.65 +/- 12.21 vs 12.2 +/- 2.7 U/L and (96.35 +/- 58.83 vs 43.2 +/- 14.5 ng/ml, respectively). In all articular compartments and in synovial fluid, the level of cytokine IL-8 exceeded concentrations of the other cytokines. In blood serum, only IL-10 levels were markedly increased as against the control group (17.35 +/- 5.82 vs 9.80 +/- 4.40 pg/ml). DISCUSSION: Primary osteoarthritis is the most common joint disease that deteriorates with age. Its symptoms are pain and a lower range of motion in the joint affected. The initial involvement of articular cartilage progresses to degenerative changes involving synovial and bony structures. This degenerative disease gradually develops into an inflammatory disease. At this stage, osseous tissue shows an increase in metabolism and bone destruction results. In the control of inflammatory reactions by the immune system, cytokines, among other proteins, play an important role: some may enhance inflammation by activating leukocytes (IL-1, TNF-alpha, IL-8) while others, such as IL-10, have anti-inflammatory effects. CONCLUSION: During osteoarthritis, the articular compartment shows high metabolic processes that, in some patients, may increase and even persist some time after arthroplasty.

Biological and biochemical properties of the carbon composite and polyethylene implant materials.

We studied the biocompatibility of the carbon composites and polyethylene materials with and without collagen or collagen and proteoglycan cover. We used the in vitro technology to study the adhesion of model cells evalution, their metabolic activity and the production of TNF-alpha as a cytokine model. Under in vivo condition, the biocompatibility of tested polymers were studied in the implantation experiment, subcutaneously in the interscapular region in the laboratory rat. We have found in the in vitro assay favorable proliferation and the smallest production of pro-inflammatory TNF-alpha cytokine in cells adherent to the hydrophobic polyethylene material coated with biological macromolecules. Using in vivo tests performed by the implantation of materials to the rat we demonstrated that the materials are not cytotoxic. The tissue capsule surrounding the implants was not significantly influenced by the type of the implant and the pre-treatment by the biological molecules. However, the foreign-body giant multinucleated cells were observed only in the vicinity of the collagen - covered hydrophobic polyethylene implant. Interestingly, while the collagen coating improved the biocompatibility of tested polymers in vitro, the inflammatory reaction against this covered materials was higher under in vivo conditions. The pre-treatment of carbon composites by both types of biological macromolecules reduced the occurrence of carbon debris in the implantation site. The tested carbon composites and polyethylene materials are not toxic. The pre-treatment of the materials by extracellular matrix components increased their biological tolerance in vitro and reduced implant wears in animal experiment, which can be important for the medical application.

[Relation between cytokines, adhesive immunoglobulins and matrix metalloproteinases in osteoarthritic joints]. / Vztahy mezi vybranými cytokiny, adhezivními imunoglobuliny a matrixovými metaloproteinázami v osteoartritickém kloubu.

BACKGROUND: The dysbalance of proteinase production and their inhibitors leads to destruction of tissues in many pathological processes. Recently it was shown that the expression of proteinases is regulated also by interactions of cells which is mediated by adhesive molecules. We wanted to find out whether this mechanism is involved also in the destruction of joints in osteoarthritis. METHODS AND RESULTS: Cartilage, synovial and subchondral bone tissues, and synovial fluids were obtained from 23 joints after total endoprosthesis surgery. The solid tissues were extracted by TRIS buffer. The investigated protein concentrations were assessed immunochemicaly. In all specimens gelatinase A, gelatinase B, stromelysin-1, TIMP-1, soluble adhesive molecules sICAM-1 and sVCAM-1 and cytokines TNF-alpha and IL-8 were found. In cartilage and synovial fluid the proteolytic potential of metalloproteinases was balanced with high concentrations of their inhibitor TIMP-1 (259.4 +/- 105.2 pmol/g protein vs 2343.8 +/- 637.5 pmol/g protein in synovial fluid, p < 0.00001, and 178.9 +/- 175.7 pmol/g dry weight vs 647.2 +/- 561.3 pmol/g dry weight in cartilage, p < 0.001) but in synovial tissues and pathological subchondral bones was not (257.4 +/- 617.2 pmol/g dry weight vs 171.3 +/- 170.8 pmol/g dry weight in synovium, p = 0.61716, and 17.4 +/- 15.4 pmol/g dry weight 33.6 +/- 33.3 pmol/g dry tissue in pathological subchondral bone, p = 0.16705). CONCLUSION: From correlation analysis ensues that the bond of ICAM-1 and VCAM-1 on chondrocytes with appropriate integrin ligands probably leads to up-regulation of gelatinase A and B and to down-regulation of TIMP-1. Moreover it is apparent that TNF-alpha up-regulates both investigated adhesive molecules followed and stromelysin-1 and TIMP-1.

[Monitoring markers of bone turnover in multiple myeloma]. / Monitorace markeru kostní prestavby u mnohocetného myelomu.

PURPOSE: To assess the efficacy of bone turnover markers monitoring for prediction of overall survival and event-free survival in patients with multiple myeloma. To determine the value of initial measurement for differential diagnosis and evaluation of myeloma stage and activity. METHODS AND RESULTS: We evaluated 79 consecutive, previously untreated patients with myeloma and 24 with monoclonal gammopathy of undetermined significance. Urine excretion rates of pyridinoline (PYR) and deoxypyridinoline (DPYR) as markers of bone resorption and serum osteocalcin (OC) as marker of bone formation were measured at diagnosis and further repeatedly during the course of disease. High-performance liquid chromatography (HPLC) was used to measure urinary excretion rates of PYR and DPYR, serum OC levels was analysed using RIA method. Significant correlation was found between PYR and DPYR levels (p < 0.001) and between DPYR and OC (p < 0.05). Significantly higher urine PYR and DPYR (p < 0.001 and p < 0.01, respectively), but not serum OC, were found in myeloma comparing MGUS. In patients with MM PYR and DPYR levels were significantly higher in stage III vs. I and II (p < 0.01), the correlation with disease activity was not found. The grade of bone involvement according to RTG (osteolysis, osteoporosis, absence of bone lesions) was not reliable as a prognostic factor in our study. Univariate overall survival analysis showed prognostic significance for initial PYR (p < 0.05) but not for DPYR and/or OC. Rapid decrease of PYR (after 1 month of chemotherapy) was associated with the shortest median survival (608 days), however, significant difference was observed only comparing the patients in which PYR decreased 12 months after start of chemotherapy. In the univariate analysis increase of DPYR was the only variable significant for event-free survival (p < 0.05), increase of PYR tend to be significant (p < 0.1). CONCLUSIONS: We confirmed possible contribution of pyridinium cross-links for differential diagnosis of MM and MGUS and the correlation with advanced stage of MM. Initial measurement of PYR and monitoring of both PYR and DPYR during the course of myeloma could be helpful for prediction of overall survival and event-free survival. According to our experience OC is not useful for diagnosis, assessment of disease stage and activity and prediction of survival of patients with MM.

Total and local changes in the arthritis adjuvans.

Arthritis adjuvans was studied in the murine model. An effect of different treatment (methotrexate, tauredon, collagen hydrolysate) was estimated in the course of developing disease (day 3, 5, 11 and 21). Repeated evaluation of body weight and peripheral blood leukograms as a total response of organism was performed. Oedema of paw periarticular and tail regions, light- and electron-microscopical screening and immunohistochemical investigation of prevalence of interleukin-1-beta (IL-1beta) and tumour necrosis factor (TNF-alpha) were estimated. The most pronounced benefit effect of methotrexate at stabilization of the monocytes blood level, synovial membrane cell invasion and TNF-alpha immunopositivity was ascertained.

Determination of pyridinium crosslinks in serum an optimization of sample preparation.

Urinary pyridinoline (UPD) and deoxypyridinoline (UDPD) are selective markers in kinetic studies of mature collagen degradation in connective tissue, especially in bone. In patients with renal dysfunction, the determination of UPD and UDPD is not entirely reliable, while in anuretic patients it is impossible. As renal dysfunction is considered a risk factor for bone diseases, it is essential to determine both markers directly in the serum (SPD and SDPD). Due to the high serum concentrations of proteins, which during acid hydrolysis are converted to amino acid hydrochlorides, the system butanol-water is sometimes separated into two phases during sample preparation. Should this fact not be taken into account, the usual sample processing on a cellulose sorbent could yield substantially lower false results. This calls for some preventive measures: to ensure the homogeneity of the system containing n-butanol it is recommended to add an appropriate third component, e.g. methanol.

[Effect of clodronate in patients with multiple myeloma. Evaluation of specific markers of bone resorption]. / Ucinek klodronátu u pacientu s mnohocetným myelomem. Hodnocení specifickými markery osteoresorpce.

BACKGROUND: In adjuvant therapy of patients with multiple myeloma among others anti-absorption properties of bisphosphonates are used. The objective of the present investigation was to evaluate the effect of long-term oral treatment route clodronate on the bone metabolism in this condition. As markers of bone reabsorption, assessment of pyridinoline and deoxypyridinoline in urine was used. METHODS AND RESULTS: We investigated in an open clinical trial 22 patients with multiple myeloma with bone changes confirmed on X-ray who had a normal calcium and creatinine serum concentration: the control group (A) comprised 13 patients (mean age 59 years, range 38-74 years), the experimental group (B) 9 patients (mean age 54 years, range 42-60 years). The patients in both groups were treated by chemotherapy, to group B concurrently clodronate 3 x 800 mg was administered by the oral route. No statistically significant differences were found between the two groups during the mean follow up period after assessment of the diagnosis (27 vs. 40 months) nor in the clinical stage of the disease. After intervals of 0, 3 and 12 months the excretion of pyridinoline and deoxypyridinoline in urine was assessed by liquid chromatography. At the same time also other parameters of osteoresorption were assessed (urinary calcium and hydroxyproline excretion). At the onset of the investigation the pyridinoline and deoxypyridinoline excretion in groups A and B did not differ statistically (pyr; 120.63 +/- 23.76 vs. 136.23 +/- 22.13 mumol/mmol creatinine. d-pyr: 19.61 +/- 3.65 vs. 22.76 +/- 5.40 mumol/mmol creatinine). After three months in group B a statistically significant drop of excretion of both metabolites was recorded (pyr. to 84.11 +/- 38.67 mumol/mmol creatinine, d-pyr. to 15.23 +/- 6.10 mumol/mmol creatinine). Their significantly reduced excretion persisted also after one year (pyr. 85.52 +/- 60.96 mumol/mmol creatinine, d-pyr. 12.6 +/- 6.56 mumol/mmol creatinine). CONCLUSIONS: Using specific markers, pyridinoline and deoxypyridinoline, the authors proved the favourable effect of long-term administration of clodronate on the bone metabolism in patients with multiple myeloma.