Hyaluronan concentration and molecular mass in psoriatic arthritis: biomarkers of disease severity, resistance to treatment, and outcome.

Objective: Low molecular mass hyaluronan causes inflammatory processes and can act as a pro-inflammatory cytokine in skin and other sites of activity in psoriatic arthritis (PsA). This study investigated whether the molecular mass distribution of hyaluronan (HA) in skin and the quantity of circulating HA are related to the clinical inflammatory picture in PsA with active disease and to the effect of treatment with anti-tumour necrosis factor-α (anti-TNF-α) adalimumab. Methods: Twenty patients with TNF-α-naïve active polyarticular PsA were included in this prospective clinical trial of treatment with 40 mg s.c. adalimumab according to standard procedure. Clinical activity, patients' assessments, and skin biopsies were captured at inclusion and at the 12 week follow-up. Ten healthy individuals were recruited for comparison of HA analyses. Histochemistry of skin inflammation, serum HA, and molecular mass of HA were determined. Results: Overall improvements in clinical parameters were observed. Eight of 18 patients reached minimum disease activity after 12 weeks and disease activity was significantly reduced (p < 0.0001). Patients with elevated serum HA values were significantly older, had later onset of arthritis and more deformed joints, still had swollen joints after treatment, and had more circulating inflammatory biomarkers. More severe disease pathology showed a wide spectrum of high-molecular-mass HA accompanied by low mass HA. The treatment appears partly to normalize the HA mass distribution. Conclusion: HA concentration and mass seem to be two possible factors in the inflammatory pathology of PsA acting as biomarkers for disease severity, resistance to treatment, and worse outcome.

Bone marrow hyaluronan and reticulin in patients with malignant disorders.

Myelofibrosis is commonly seen in patients with chronic myeloproliferative diseases and sometimes in myelodysplastic syndrome, acute leukaemia and lymphoproliferative diseases. The fibrotic process is evaluated by grading the amount of collagen deposited in the bone marrow interstitium. The established method to evaluate bone marrow fibrosis is staining for reticulin to visualise the collagen fibres. However, the extra cellular matrix does not only contain collagens but also other components, e.g. glycosaminoglycans of which hyaluronan is the most abundant. Hyaluronan is important for structural and cellular functions. Earlier studies have shown that there is a positive correlation between hyaluronan and reticulin staining in healthy volunteers and in patients with de novo acute myeloid leukaemia. In this study bone marrow biopsies from 43 patients with a malignant disease involving the bone marrow were compared with 18 patients with a malignant disease not involving the bone marrow. The intensity of hyaluronan grading was significantly higher in the patients with disease involving the bone marrow compared to the healthy controls but not compared to the patients without disease involving the bone marrow. The staining intensity of reticulin in the bone marrow was significantly higher in the patients with disease involving the bone marrow, compared to those without disease involving the bone marrow and to the controls. In all patients and the controls there was a correlation between hyaluronan and reticulin.

The stromal reaction in basal cell carcinomas. A prerequisite for tumour progression and treatment strategy.

Specimens of basal cell carcinomas collected from 28 patients were classified into three groups: superficial, nodular, and infiltrative, according to their microarchitecture. The specimens were then subjected to histological characterization by means of a biotinylated hyaluronan-binding probe (HABP). By using Ki-67 and PCNA the proliferative activity of the BCC tumours was evaluated with immunohistological techniques. In superficial BCC the tumour islands displayed moderate hyaluronan (HA) staining. Feeble proliferation, denoted by modest mitotic activity and weak Ki-67 and PCNA immunoreactivity, occurred within the tumour islands. The surrounding connective tissue resembled normal skin, and no differentiated tumour stroma was observed. In nodular BCC, the HA staining of the tumour strands was weak to moderate, denoting increased proliferative activity. The differentiated surrounding tumour stroma stained strongly for HA. Tumour islands of infiltrative BCC stained weakly to moderately to HA and evidenced intense proliferation. The intensely HA-stained tumour stroma ended abruptly and the adjacent areas were almost devoid of HA. This study showed that the proliferative activity of BCC cells is associated with increased expression of HA in the tumour stroma. Modification of tumour-associated connective tissue indicates a close relationship between the tumour cells and the adjacent matrix. In particular, in infiltrative BCC, such alterations include degeneration and possible modification and remodelling of the surrounding extracellular matrix. These processes involving areas of probable importance for tumour progression, should be considered when deciding the extent of intended surgical resection.

Hyaluronan in joint disease.

Hyaluronan is a major component of synovial fluid. It is synthesized in the joint and partly degraded in joint capsule and partly carried by lymph to lymph nodes and the general circulation. During various joint disorders, especially inflammatory ones, the amount of hyaluronan in the joint and its turnover rate are increased leading to pathologically high serum levels of the polysaccharide. These levels are a complex function of synovitis mass and the physical activity of the patient. Intra-articular injection of hyaluronan has been used in the treatment of osteoarthritis. Although it results in a relief of pain, controlled clinical trials have not yet convincingly shown any beneficiary effects of the treatment.

Availability of iron and degree of inflammation modifies the response to recombinant human erythropoietin when treating anemia of chronic disease in patients with rheumatoid arthritis.

Forty-six patients with rheumatoid arthritis (RA) and documented anemia of chronic disease (Hb < 100/110 g/l) were randomized to receive either human recombinant erythropoietin (r-HuEPO, n = 36, 300 U/kg body weight) or placebo (n = 10) for 12 weeks in a multicenter study. An adequate response was defined as elevation of Hb > or = 120 g/l. Relevant clinical and laboratory assessments were made to evaluate efficacy and secure safety. A significant elevation in Hb from week 10 onwards was noted in twenty-six patients (five drop-outs) out of nine patients receiving placebo (one drop-out) (12 +/- 1.2 g/l vs 4 +/- 0.5 g/l; Hb elevation from 95 g/l to 107 g/l vs 93 g/l to 97 g/l, P < 0.05). Only 14.6%, however, were considered responders according to preset criteria. In the responders a lower initial CRP, a significant reduction in ESR but not in CRP was seen compared to the remaining r-HuEPO group. A significant elevation of energy level was noted in the r-HuEPO group; otherwise, no differences in clinical variables were seen. No serious adverse effects were noted. When analyzing patients receiving oral iron in combination with r-HuEPO and adding five additional, openly selected patients receiving both adequate iron supplementation and r-HuEPO, there was a significant weekly elevation of Hb from week 8 onwards in favor of combination therapy over the ones only receiving r-HuEPO (18 +/- 1.1 g/l vs 7 +/- 1.1 g/l, P < 0.05). The initial six responders had now reached ten of whom seven belonged to the combination therapy group. Response to r-HuEPO in RA patients appears to be dependent on availability of iron and on the degree of inflammation. If r-HuEPO treatment is considered, iron deficiency should always be corrected and strenuous efforts should have been made to control the disease itself.

Hyaluronan in human deciduous tooth germs in the bell stage. Histochemistry and immunohistochemistry.

The primary aim of the present study was a localization of hyaluronan (HA) in human deciduous tooth germs in the bell stage. HA was compared to the content of chondroitin sulfates (CSs). HA was detected with a biotin-labeled HA-binding protein (HABP) and CS with a monoclonal antibody. As controls, enzyme digestions were carried out. Furthermore, the glycosaminoglycans were investigated histochemically with enzyme digestions followed by alcian blue staining. The investigation showed a considerable content of HA in the stellate reticulum, although CS was also found, primarily when treatment with protease was omitted. The dental papilla contained both HA and CS, while the predentin and the dentin contained only CS. The enamel did not contain any CS, but some staining with HABP was observed along the borderline between the ameloblasts and the enamel. The significance of HA in the stellate reticulum is discussed. The importance of carrying out investigations with and without protease digestions is stressed.

Localization and quantity of hyaluronan in urogenital organs of male and female rats.

The histochemical distribution of hyaluronan was analysed in various urogenital organs of male and female (non-pregnant and pregnant) rats by use of a hyaluronan-binding protein and avidin biotin/peroxidase staining. Microwave-aided fixation was used to preserve the extracellular location of hyaluronan. The concentrations of hyaluronan in the different tissues were measured with a highly sensitive radio-assay. Hyaluronan accumulated predominantly in the connective tissue around smooth muscle fibres and in the subepithelial lamina propria. Abundant hyaluronan also occurred in perivascular and perineural connective tissue. In the female urogenital organs, hyaluronan content was high in the vagina and urinary bladder, and highest in the vagina during pregnancy. In the uterus, the surface epithelium of the endometrium stained intensely. In the ovary, the zona pellucida of the oocyte and the theca interna cell layer of the follicles and the follicular fluid of mature follicles exhibited prominent staining. The corpus luteum was devoid of hyaluronan, whereas enlarged corpora lutea of pregnancy exhibited weak, patchy staining. In male urogenital organs, staining for hyaluronan was absent from the testis and epididymis, whereas the erectile connective tissue of the penis stained intensely. The hyaluronan concentrations were high in penile tissue and urinary bladder, while testis, epididymis and the ductus deferens contained only little hyaluronan.

The Fibromyalgia Impact Questionnaire, a Swedish translation of a new tool for evaluation of the fibromyalgia patient.

The American Fibromyalgia Impact Questionnaire (FIQ) was translated into Swedish. The FIQ is a brief 10-item self-administered instrument designed to evaluate the special problems bothering fibromyalgia patients. After the translation procedure, 73 Swedish women fibromyalgia patients answered the translated FIQ. The questionnaire was fast and simple to handle both for the patients and the investigators. There was a good level of correspondence between FIQ items and related items in the Health Assessment Questionnaire, the Nottingham Health Profile and the Psychological General Well-Being Index. There were no significant differences in a test-retest analysis. The study has provided evidence that the translated and slightly modified Swedish version of the American Fibromyalgia Impact Questionnaire has the validity and sensitivity to be able to function as a relevant and comprehensible tool to monitor FMS-patients.

Local anti-type II collagen antibody production in rheumatoid arthritis synovial fluid. Evidence for an HLA-DR4-restricted IgG response.

OBJECTIVE: To investigate the production of type II collagen (CII) antibodies in the synovial fluid (SF) of rheumatoid arthritis (RA) patients, and to examine the HLA dependence of this local production. METHODS: The ELISPOT method was used for enumerating anti-CII-reactive cells. Serologic tissue typing was performed. RESULTS: Anti-CII-reactive cells were found in the SF of 16 of 31 patients, but not in any of the peripheral blood samples obtained in parallel. SF anti-CII antibody production showed no correlation with clinical parameters, but its frequency increased significantly with age. The IgG anti-CII response occurred exclusively in patients who were positive for HLA-DR4 and was significantly associated with DR4. CONCLUSION: Anti-CII production may be important in local immune complex formation. The indirect demonstration of a DR4-restricted T cell response to CII is an indication of a pathogenetic role of collagen autoimmunity in RA.

Serum hyaluronan and aminoterminal propeptide of type III procollagen in primary biliary cirrhosis: relation to clinical symptoms, liver histopathology and outcome.

Hyaluronan (HA) and aminoterminal propeptide of type III procollagen (PIIINP), two biochemical connective tissue markers, were determined in 76 patients with primary biliary cirrhosis (PBC). The HA and PIIINP concentrations were significantly increased compared with controls (P less than 0.001). Both HA and PIIINP levels correlated significantly with conventional liver-function tests. All patients with stage IV PBC showed increased concentrations of both these variables. However, HA was a better marker with regard to prediction of development of cirrhosis as well as prediction of symptoms. Furthermore, HA also showed a negative correlation with time of survival (P less than 0.05). The present data indicate that HA is a more sensitive marker of liver damage in PBC than PIIINP.

Catabolism of hyaluronan in the knee joint of the rabbit.

Catabolism of hyaluronan was studied by injecting hyaluronan labelled with [125I]-tyramine cellobiose ([125I]-TC) into knee joints of rabbits. After endocytosis [125I]-TC remains intracellularly allowing localization of the site of catabolism. At 6 hours after injection 63% could be recovered in and around the joint, while at 48 hours 32% remained locally. Chromatography showed that 12% of the injected tracer was degraded in joint tissues at 6 hours, increasing to 33% at 24 hours. There was no apparent degradation within the joint fluid. No tracer was found in the regional lymph glands, but 16% of the injected tracer was detected in the liver at 24 hours. This investigation demonstrates that hyaluronan in the joint can be degraded both locally and in the liver.

Hyaluronan in human skeletal muscle of lower extremity: concentration, distribution, and effect of exercise.

The concentration and localization of hyaluronan (HYA) were determined in biopsy specimens from resting human quadriceps femoris and anterior tibial muscles. The influence of physical exercise on HYA concentrations in the quadriceps femoris muscle and in blood was also evaluated. A sensitive radioassay was used for the quantification of HYA. The distribution of the glycosaminoglycan was demonstrated using a histochemical method that involved microwave-aided fixation and an HYA-binding protein. At rest, the muscle HYA concentration was 34.9 +/- 23.6 (SD) micrograms/g muscle wet wt with a large interindividual variation. Exercise had no significant effect on the muscle HYA concentration. The serum HYA concentration increased from 35.9 +/- 22.7 to 53.4 +/- 57.1 micrograms/l during exercise, but 30 min after the exercise the HYA concentration was significantly lower (19.1 +/- 6.3 micrograms/l) than the initial preexercise value. In resting skeletal muscles of the lower extremity, HYA was heterogeneously distributed in the perimysium and endomysium. Perivascular and perineural connective tissues were distinctly HYA positive.

Serum concentrations of hyaluronan and proteoglycan in joint disease. Lack of association.

Circulating hyaluronan originating from the synovial membrane and circulating proteoglycan released from cartilage were determined by specific assays in patients with osteoarthritis (OA), rheumatoid arthritis (RA), reactive arthritis or juvenile chronic arthritis (JCA). Elevated hyaluronan concentrations were found in OA and RA, suggesting proliferation of the synovial membrane in both diseases. The proteoglycan concentrations were highest in OA and polyarticular JCA indicating increased turnover of cartilage matrix. The concentrations of the macromolecules did not correlate except in the group with JCA. Serum concentrations of hyaluronan and proteoglycan thus differ between disease groups and may reflect different aspects of the arthritic process.

Hyaluronan in pleural effusions and in serum.

It has been suggested that a high level of hyaluronan (hyaluronic acid, HYA) in pleural fluid is an indicator of malignant mesothelioma. In 78 consecutive patients with pleural effusion of various causes the HYA concentration was measured in pleural fluid samples and in serum. Nine patients had malignant pleural mesothelioma, and in three of them the HYA level in pleural fluid was 100 mg/l or more. In 42 patients with effusions due to metastatic malignancy, the mean HYA in the pleural fluid was 75 mg/l, and in five the HYA level was above 100 mg/l. Cardiac insufficiency caused the effusion in 11 patients, of whom two had a level above 100 mg/l in pleural fluid. Four patients had a serologically confirmed viral infection and had HYA levels in pleural fluid of 8, 157, 335, and 554 mg/l, respectively. One patient had postinfectious effusion with an HYA level in pleural exudate of 748 mg/l, the highest in this investigation. Two patients had benign asbestos pleural effusions, and both had high pleural HYA levels (256 and 490 mg/l, respectively). The serum HYA values were much lower than in the pleural fluid, namely from 15 to 480 micrograms/l; the levels were independent of the levels in the pleural fluid. Thus, a high level of HYA in pleural fluid is not specific for mesothelioma but can occur in other malignant or benign diseases, and a low level does not exclude mesothelioma.

Localization of hyaluronan in various muscular tissues. A morphological study in the rat.

The histochemical distribution of hyaluronan (hyaluronic acid, HYA) was analysed in various types of muscles in the rat by use of a hyaluronan-binding protein (HABP) and the avidin-biotin/peroxidase complex staining procedure. Microwave-aided fixation was used to retain the extracellular location of the glycosaminoglycan. In skeletal muscles, HYA was detected in the connective tissue sheath surrounding the muscles (epimysium), in the septa subdividing the muscle fibre bundles (perimysium) and in the connective tissue surrounding each muscle fibre (endomysium). HYA was heterogeneously distributed in all striated muscles. In skeletal muscles with small fibre dimensions (e.g., the lateral rectus muscle of the eye and the middle ear muscles), HYA was predominantly accumulated around the individual muscle fibres. Perivascular and perineural connective tissue formations were distinctly HYA-positive. In cardiac muscles, HYA was randomly distributed around the branching and interconnecting muscle fibres. In comparison, smooth muscle tissue was devoid of HYA.

Circulating hyaluronan in the myelofibrosis/osteomyelosclerosis syndrome and other myeloproliferative disorders.

The serum concentration of hyaluronan (HYA) was determined in 59 patients with various myeloproliferative disorders, including 33 patients with idiopathic myelofibrosis. In 18 patients the serum concentration of the aminoterminal propeptide of type III procollagen (PIIINP) was measured concomitantly. Raised serum HYA levels were seen in patients with active disease compared with age-matched healthy subjects, whereas no significant difference in serum HYA was seen between patients with stable disease and age-matched controls. Serum HYA concentrations correlated significantly with the leukocyte count (rho = 0.38; P less than 0.02) and with the serum concentration of PIIINP (rho = 0.50; P less than 0.001). During cytotoxic treatment, the serum HYA and PIIINP concentrations decreased in concert with declining leukocyte counts. These findings suggest that clonal expansion is accompanied by a bone marrow stromal reaction similar to the repair processes following injury to soft connective tissues. The relatively modest changes in serum HYA with frequent overlaps between patient categories and healthy subjects imply that the clinical utility of single determinations of serum HYA in the present disease groups is restrained. On the other hand, sequential measurements of HYA may provide a reflection of the myeloproliferative process in individual patients.