Spectroscopic assessment of endogenous porphyrins in a rheumatoid arthritis rabbit model after the application of ALA and ALA-Me.

The sensitization of inflamed knee tissues with endogenous porphyrins was studied by means of fluorescence spectroscopy in an experimental model of rabbit rheumatoid monoarthritis after intraarticular (i.a.) or intravenous (i.v.) injections of 5-aminolevulinic acid (ALA) or ALA methyl ester (ALA-Me). Fluorescence measurements in vivo on the skin of the inflamed knee joint showed the dominance of protoporphyrin IX (PpIX). The highest fluorescence intensity was registered 2h after i.a. injection of ALA and ALA-Me. Comparative analysis of the PpIX fluorescence spectra ex vivo revealed that more PpIX accumulated in the tissues of the inflamed joint than in the tissues of the control joint, and that ALA-Me induced about five times more PpIX in the inflamed synovium than ALA. Meanwhile, the cartilages of the inflamed and control knee joints also accumulated water-soluble porphyrins. Thus, in vivo and ex vivo spectroscopic assessment of endogenous porphyrins in rabbit rheumatoid arthritis tissues implied that the injection of ALA is more appropriate for the diagnostics of inflammation due to the higher PpIX fluorescence intensity on the skin surface, while ALA-Me is more appropriate for the therapeutic applications due to the higher and more selective accumulation of PpIX in the inflamed synovium.

[Hemostasis in patients with rheumatoid arthritis].

AIM: To study hemostasis with reference to rheumatoid arthritis (RA) activity, extraarticular manifestations and vascular damage. MATERIAL AND METHODS: We studied hemostasis in 104 RA patients. The following parameters were tested: partial thromboplastin time activation (PTTA), fibrinolytic activity (FA), prothrombin index, lupus anticoagulant (LA), mean platelet volume (PV), antibodies against cardiolipin (aCL) and antibodies against beta2-glycoprotein 1 (alphabeta2-GP1). The disease activity index was estimated by DAS-28 including tender and swelling joints count, erythrocyte sedimentation rate and disease global activity rated by the patient. RESULTS: We found out an increased number of platelets in 54.4%, prolongation of FA and PTTA in 72.5 and 12.5% patients. The rest blood coagulation tests were normal in most the examinees. The LA antibodies were absent, while aCL and alphabeta2-GP1 antibodies were identified in 10.6 and 14.4%, respectively. The number of platelets increased while hemoglobin decreased in relation to higher disease activity and PV was significantly lower in patients with extraarticular manifestations. No significant influence of vascular damage or comorbidities on hemostasis was found. CONCLUSION: No relation was established between coagulation factors and the disease activity except for increased number of platelets in patients with higher activity and lower PV in patients with extraarticular manifestations. However, patients with aCL and alphabeta2-GP1 need constant monitoring because of the risk to develop thrombosis.

[Antineutrophil cytoplasmic antibodies in patients with diffuse diseases of connective tissue]. / Antineitrofil'nye tsitoplazmaticheskie antitela u bol'nykh s diffuznymi bolezniami soedinitel'noi tkani.

AIM: To investigate the significance of antibodies against neutrophil cytoplasmic antigens (ANCA) in patients with rheumatoid arthritis (RA), systemic lupus erythematosus (SLE) and systemic sclerosis (SS), their relations with the syndromes and laboratory indices. MATERIAL AND METHODS: The sera from 277 patients suffering from connective tissue disease and 31 healthy persons were examined. Of patients, 103, 126 and 48 had RA, SLE and SS, respectively. The patients in each group were subdivided into ANCA positive (ANCA+) and ANCA negative (ANCA-). The patients were matched within the groups by age, sex and disease duration. There were 41 such pairs in RA group, 23 in SLE and 13 in SS group. Questionnaires and laboratory tests (ANA, RF, a-DNA, a-MPO, a-Scl-70, a-PR3, a-CL) were used in the examination. RESULTS: The sensitivity of ANCA in SLE patients group was as high as 58.7%, specificity--93.5%. In other groups ANCA were less frequent. ANCA were significantly associated with skin vasculitis and ANA prevalence but the disease activity in SLE was not related to this feature. Anemia and antibodies against cardiolipin were found significantly more frequently in ANCA positive RA group. In SS group the inverted clinical association with kidney damage was seen but a-DNA were more prominent in ANCA+ group. Subspecificity for a-MPO and a-PR3 and lactoferin by ELISA were revealed less often than a-ANCA by immunofluorescence. Only two SLE patients with a-lactoferin antibodies were evidently different in prognosis while the other ones did not differ in the disease course within their group. CONCLUSION: The ANCA pattern in connective tissue diseases is found rather often but only few clinical and laboratory associations could be established such as skin vasculitis and ANA domination in SLE group, anemia and a-CL in RA group and a-DNA in SS ANCA+ groups. The validity of ANCA test is not so significant as it is in vasculitis patients.

Amelioration of collagen-induced arthritis by thrombin inhibition.

The deleterious role of fibrin deposition in arthritic joints prompted us to explore the effect of the thrombin inhibition on the course of collagen-induced arthritis (CIA) in the mouse. CIA was induced in male DBA/1J mice using native chicken type II collagen. The thrombin inhibitor polyethyleneglycol-hirudin (PEG-hirudin) was given for 16 days, starting 20 days after the first immunization (preventive treatment) or at the onset of clinical signs of arthritis (curative treatment). All the mice treated with PEG-hirudin had a significantly prolonged clotting time compared with control mice. PEG-hirudin, administered in a preventive way, led to significantly reduced incidence and severity of CIA during most of the treatment period, as assessed by clinical scoring. Accordingly, histological features showed a significant diminution of synovial hyperplasia in PEG-hirudin-treated mice compared with untreated mice. There was also a significant downmodulation of the synovial proinflammatory IL-1beta and IL-12p35 cytokine mRNAs in treated mice. Intra-articular fibrin, evaluated by immunohistochemistry, was significantly reduced in treated mice compared with control mice and correlated with both clinical and histological scorings. Most importantly, once arthritis was established, PEG-hirudin also showed a curative effect. In conclusion, PEG-hirudin can both prevent the onset of CIA in a dose-dependent manner and ameliorate established arthritis, suggesting that thrombin inhibition may offer a new therapeutic approach in arthritis.