Monitoring periodontal disease status in smokers and nonsmokers using a gingival crevicular fluid matrix metalloproteinase-8-specific chair-side test.

BACKGROUND AND OBJECTIVE: With current periodontal diagnostic tools it is difficult to identify susceptible individuals or sites at risk. The aim of this study was to evaluate the efficacy of the matrix metalloproteinase (MMP)-8-specific chair-side dip-stick test in longitudinally monitoring the periodontal status of smoking (S) and nonsmoking (NS) patients with chronic periodontitis, using their gingival crevicular fluid (GCF) MMP-8 concentrations. MATERIAL AND METHODS: Clinical parameters, MMP-8 test results and concentrations were monitored in 16 patients after initial treatment and in 15 patients after scaling and root planing (SRP), every other month, over a 12-mo time period. Progressing and stable sites, and sites with exceptionally high MMP-8 concentrations, were analysed in smokers and nonsmokers. RESULTS: SRP reduced the mean GCF MMP-8 levels, test scores, probing depth (PD), attachment loss (AL) and bleeding on probing (BOP). In sites of periodontal disease progression, the distribution of MMP-8 concentrations was broader than in stable sites, indicating a tendency for elevated concentrations in patients with periodontal disease. The mean MMP-8 concentrations in smokers were lower than in nonsmokers, but in smokers' and nonsmokers' sites with progressive disease, MMP-8 concentrations were similar. Sites with exceptionally elevated MMP-8 concentrations were clustered in smokers who also showed a poor response to SRP. In these sites, the MMP-8 concentration did not decrease with SRP and these sites were easily identified by the MMP-8 test. CONCLUSION: Persistently elevated GCF MMP-8 concentrations may indicate sites at risk, as well as patients with poor response to conventional periodontal treatment (e.g. SRP). MMP-8 testing may be useful as an adjunct to traditional periodontal diagnostic methods during the maintenance phase.

Changes in gingival crevicular fluid matrix metalloproteinase-8 levels during periodontal treatment and maintenance.

OBJECTIVES: The aim of this study was to investigate the effect of scaling and root planing (SRP) and the maintenance phase of treatment on the gingival crevicular fluid (GCF) matrix metalloproteinase-8 (MMP-8) levels. MATERIALS AND METHODS: Clinical measurements and GCF samples were taken from four sites in 20 adult periodontitis patients before and after SRP and during a 3-month maintenance phase of treatment. MMP-8 levels were measured from GCF samples by time-resolved immunofluorometric assay (IFMA) with monoclonal antibodies. RESULTS: SRP improved the clinical indices as would be predicted, 6.1 mm (SD = 1.4) at baseline compared with 4.3 mm (SD = 1.6) post-treatment (P < 0.001). Attachment level (AL) reduced but not significantly between these two visits 13.4 mm (SD = 2.4) compared with 12.8 mm (SD = 2.4) (P < 0.08) post therapy. GCF MMP-8 levels reduced after initial treatment from 33.8 micro g/30 s sample to 23.5 micro g/30 s, which just failed to reach statistical significance (P = 0.07). However, when MMP-8 levels were expressed as a concentration, the differences following initial therapy were significant (54.1 ng/ micro L at baseline compared with 34.2 ng/micro L post treatment; P < 0.005). The difference, however, between the baseline MMP-8 levels (33.8 ng/30 s) and the final visit (16 ng/30 s) following maintenance was markedly significant (P < 0.001) for both absolute amounts and on a concentration basis. CONCLUSION: In conclusion, clinical improvement following SRP was associated with significant reductions in MMP-8 levels. The GCF concentration of MMP-8 decreased after initial therapy but reduced even more dramatically (approximately 50%) following a 3-month period of maintenance (P < 0.001).

Scientific basis of a matrix metalloproteinase-8 specific chair-side test for monitoring periodontal and peri-implant health and disease.

Matrix metalloproteinases (MMPs), especially collagenase-2 (MMP-8), are key mediators of irreversible tissue destruction associated with periodontitis and peri-implantitis. MMP-8 is known to exist in elevated amounts and in active form in the gingival crevicular fluid (GCF) and peri-implant sulcular fluid (PISF) from progressing periodontitis and peri-implantitis lesions and sites, respectively. (Sorsa et al. Ann. N.Y. Acad. Sci. 737: 112-131 [1994]; Teronen et al. J. Dent. Res. 76: 1529-1537 [1997]). We have developed monoclonal antibodies to MMP-8 (Hanemaaijer et al. J. Biol. Chem. 272: 31504-31509 [1997]) that can be used in a chair-side dipstick test to monitor the course and treatment of periodontitis and peri-implantitis. Monoclonal and polyclonal antibody tests for MMP-8 coincided with the classical functional collagenase activity test from GCF and PISF (Sorsa et al. J. Periodont. Res. 22: 386-393 [1988]) in periodontal and peri-implant health and disease. In future a chair-side functional and/or immunological MMP-test can be useful to diagnose and monitor periodontal and peri-implant disease and health.

Total and lipid-bound serum sialic acid in benign and malignant breast disease.

Elevation in the total sialic acid (TSA), TSA/total protein (TSA/TP) and lipid-bound sialic acid (LASA) concentration in serum occurs in breast cancer and we have studied the applicability of the assays in classification of undefined breast tumors. Sialic acid was determined by HPLC and the statistical evaluation included the receiver operating characteristic (ROC) and Youden's index analyses. In cancer patients, the serum LASA and TSA concentration was significantly higher (p < 0.05) than in patients with benign breast disease and all the markers were significantly higher (p < 0.0001) than in normal controls. All the markers had a low accuracy (AUCs < 0.75) in differentiating between breast cancer and benign breast disease and at the specificity level of 0.95 the corresponding sensitivities were 0.32 (TSA), 0.14 (TSA/TP) and 0.23 (LASA). The results indicate that both breast cancer and benign breast disease cause elevation of TSA, TSA/TP and LASA values in serum and do not provide reliable classification of undefined breast tumors.

Hyperamylasemia after cardiac surgery in infants and children.

OBJECTIVE: This study was conducted to clarify the incidence of hyperamylasemia after cardiac surgery in infants and children. DESIGN AND PATIENTS: 186 infants and children operated on at Children's Hospital. Helsinki, during an 11-month period were enrolled in the study. Serum samples were taken before and on 3 consecutive days after cardiac surgery at the intensive care unit and before discharge from the hospital. MEASUREMENTS: We measured serum total amylase and serum pancreatic amylase with two different assays (1) reduction of salivary amylase from total amylase activity and (2) measurement of mass concentration with monoclonal antibodies. RESULTS: Preoperative values for both total amylase and pancreatic isoenzymes were strongly age-related. At least one of the three tests showed postoperative hyperamylasemia (> +/- 2 SD above starting values of the age group and maximal value > 3 times the individual starting value) in 64/186 (34%) patients. 22/186 (12%) patients had abnormal results in all assays. A more than tenfold rise in pancreatic amylase, suggesting pancreatitis, was found in 14 patients (8%). Mortality was 21% in this subgroup, but 5% in the rest of the patients. Hyperamylasemia was more common after 1 year of age, and after open-heart surgery, especially homograft implantation or cardiac transplantation. CONCLUSIONS: Hyperamylasemia is a common finding after cardiac surgery in pediatric patients. Amylase isoenzyme measurements are needed for clinical decision making. Age-group-related reference values are mandatory for the right interpretation of amylase values.

Urinary trypsinogen-2 test strip for acute pancreatitis.

BACKGROUND: A simple, rapid test is specific and sensitive enough to distinguish, in patients with clinically suspected acute pancreatitis, those whose abdominal pain is indeed of pancreatic origin has proved elusive. METHODS: In two consecutive series of surgical patients in a teaching hospital, whose acute abdominal pain turned out to be due to acute pancreatitis (n-57) or extrapancreatic in origin (n=40), we studied urinary trypsinogen-2 in two ways. A test strip, incorporating monoclonal antibodies to two epitopes on trypsinogen-2, recorded a blue line when concentrations exceeded 50 microgram/L; we also measured trypsinogen-2 concentrations in the laboratory. FINDINGS: In the patients with acute pancreatitis the test strip was positive in 52 and negative in five, whereas in the 40 extrapancreatic controls there were four false positives. In a further set of 57 orthopaedic controls, one urine was strip-test positive. Concentrations of urinary trypsinogen-2 and the test-strip results were in good agreement and in only three of the 154 patients were the two approaches discrepant, at the 50 microgram/L cut-off. INTERPRETATION: These findings, in patients whose acute abdominal pain was known to be pancreatic in origin or not, are encouraging but need to be confirmed in a consecutive series of patients in whom the diagnosis of pancreatitis is in doubt.

Ultrafiltration devices tested for use in a free thyroxine assay validated by comparison with equilibrium dialysis.

A new ultrafiltration method for measuring serum free thyroxine (FT4) is presented. The method makes use of disposable ultrafiltration devices, which were selected on grounds of their ability to produce protein-free serum ultrafiltrates necessary for accurate determination of FT4. Ultrafiltrate thyroxine was measured by radio-immunoassay. As measured by this method, the mean serum FT4 concentration in reference subjects (n = 17) was 25.4 pmol/l (SD 7.8). Two patient groups were studied: hyperthyroid (n = 20, mean serum FT4 138 pmol/l, SD 69) and hypothyroid (n = 18, mean serum FT4 14.8 pmol/l, SD 6.5). These results were compared with serum FT4 concentrations, as measured by an ultrafiltration method based on the use of dialysis-tubing bags (r = 0.97), and by an equilibrium dialysis method (r = 0.95). The mean level of FT4, as measured by the ultrafiltration methodologies, was about twice as high as that measured by equilibrium dialysis. This new FT4 method is more practical than earlier ultrafiltration methods and, theoretically, analytically more accurate than equilibrium dialysis methods.

Clinical evaluation of new serum tumor markers CA M26 and CA M29 in patients with primary breast cancer.

Serum CA M26 and CA M29 values were determined in 125 women: 46 patients with primary breast cancer, 49 patients with benign breast disease, and 30 controls. The mean CA M26 and CA M29 values in breast cancer patients were higher than in patients with benign breast disease and in control subjects. When we set the cut-off level at 40 kU/1 for CA M26, 15/46 (33%) of all breast cancer patients, 11/20 (55%) of breast cancer patients with axillary node involvement, 1/1 breast cancer patient with distant metastases, 4/49 (8%) of patients with benign breast disease, and 7/30 (23%) of control subjects were above this cut-off level. For comparison, at the cut-off level of 10 kU/1 the CA M29 test was positive in 30/46 (65%) cancer cases, in 16/20 (80%) of cancer patients with nodal involvement, in 9/49 (18%) of the patients with benign breast disease, and in 10/30 (33%) of control subjects. Patients with axillary nodal metastasis showed higher values than patients without metastasis in both tests. When we used a cut-off level of 15 kU/1 for CA M29, 24% (11/46) of all breast cancer patients, 35% (7/20) of breast cancer patients with axillary node involvement, 4% (2/49) of patients with benign breast disease, and 0% of control subjects were above this cut-off level. The combination of CA M26 at the 40 kU/l cut-off level and CA M29 at the 10 kU/l cut-off level reached the diagnostic sensitivity of 0.65, specificity of 0.78, and efficiency of 0.72. In breast cancer detection among patients with breast problems serum marker CA M29 reached the sensitivity of 0.65 at the specificity level of 0.82. Our results suggest that the CA M29 marker alone is as good as the panel of two markers (CA M26, CA M29) in breast cancer diagnostics. Thus it seems that CA M29 is a promising serum tumor marker in breast cancer.

New ultrafiltration method for free thyroxin compared with equilibrium dialysis in patients with thyroid dysfunction and nonthyroidal illness.

This new ultrafiltration method for free thyroxin in serum [FT4(U)] is based on radioimmunoassay of the free hormone fraction in ultrafiltrates obtained by centrifuging serum samples in Unisep Ultracent-10 ultrafiltration devices. We compared the results obtained with those by an equilibrium dialysis method [FT4(D)]. In 36 euthyroid healthy subjects, the mean FT4(U) concentration was 24.2 pmol/L and the mean FT4(D) concentration 14.8 pmol/L. In hyperthyroid and hypothyroid patients, results by the ultrafiltration method were also approximately twice as high as those obtained by the dialysis method. In 23 patients with various nonthyroidal illnesses, mean FT4(U) was 41.2 pmol/L and mean FT4(D) 19.8 pmol/L. The mean FT4(U)/FT4(D) ratio in patients with nonthyroidal illnesses (1.97) was not significantly higher than in control subjects (1.68), making it unlikely that the increase in serum FT4 is caused by weakly protein-bound and therefore dialyzable inhibitors of thyroxin binding to carrier proteins. However, two nonthyroidally ill patients with a clearly increased FT4(U) but a normal FT4(D) concentration might have had such inhibitors, whereas for two other nonthyroidally ill patients a high molar ratio of free fatty acids to albumin is a more likely explanation for increased FT4(U) and FT4(D) concentrations. On theoretical grounds, we consider the FT4(U) concentrations analytically more nearly accurate than FT4(D) values for all patient groups studied.

Efficient test for cancer antigens: decreased levels of cancer antigen in serum after excision of breast tumor.

Monoclonal antibody technology has developed new serum marker tests which may offer advantages in the follow up of cancer patients. In breast cancer detection among symptomatic patients MCA and CA 15-3 monoclonals at best reach a sensitivity of about 0.45 at a specificity level of 0.90. We have tested these monoclonal tests (MCA and CA 15-3) by applying them to detect the postoperative decrease of marker levels in the serum among women with breast cancer or benign breast disease. The CA 15-3 postoperative decrease test (5% decrease) reached slightly higher diagnostic sensitivity (0.70) and efficiency (0.83) than the MCA postoperative decrease test (sensitivity 0.65, efficiency 0.80). The combined MCA and CA 15-3 decrease tests had a sensitivity of 0.85 and an efficiency of 0.90 at 5% decrease level. The postoperative decrease test is far superior to normal marker tests in breast cancer detection, especially when several sensitive markers are used. A 35% or larger decrease in serum levels of MCA or CA 15-3 after surgery was 100% specific for breast cancer, and detected 43% of all cancer cases. Although such a test cannot be used for screening, it could be valuable in special diagnostic situations.

Association between increased concentrations of free thyroxine and unsaturated free fatty acids in non-thyroidal illnesses: role of albumin.

We measured the concentrations of non-esterified free fatty acids and free and total thyroid hormones in serum from patients with various non-thyroidal illnesses (NTI) and chronic renal failure (CRF). The total concentration of free fatty acids was measured enzymatically and the eight most abundant fractions were determined by gas-liquid chromatography. The concentration of total free fatty acids was significantly increased in the NTI group as compared with controls (p less than 0.01); the concentrations of oleic, linoleic and linolenic acid were increased more than those of the other fractions. In NTI the serum-free thyroxine (FT4) concentration was increased (p less than 0.01) and the free triiodothyronine (FT3) concentration was decreased (p less than 0.001); these concentrations were measured by equilibrium dialysis. There was a significant correlation between the levels of total free fatty acids and FT4 in the NTI (n = 43) group (r = 0.45, p less than 0.01), and also between the levels of linoleic acid and FT4 (r = 0.35, p less than 0.05). The serum albumin concentration was decreased in the NTI group, and when free fatty acid to albumin molar ratios were calculated stronger correlations with FT4 were observed (total free fatty acids: r = 0.55; p less than 0.001; oleic acid: r = 0.30, p less than 0.05; linoleic acid: r = 0.46, p less than 0.01; linolenic acid: r = 0.35, p less than 0.05). There was no correlation between FT4 and unsaturated FFA concentrations in CRF patients, who had normal mean FT4 and total FFA levels. These results support the hypothesis that unsaturated fatty acids are involved in the increase of serum FT4 in NTI, especially when albumin levels are low.

Cancer-associated antigen CA 15-3 in the diagnostics of breast tumours.

The serum concentration of the new marker CA 15-3 was determined by a kit method, which is based on the use of two different monoclonal antibodies 115D8 and DF3, in a coated tube immunoradiometric technique. The mean CA 15-3 values in breast cancer patients (n = 40) were significantly higher than in patients with benign breast disease (n = 52, p less than 0.001) and in control subjects (n = 32, p less than 0.001). When we used the cut-off level 35 kU/l for CA 15-3, 0/32 of control subjects, 1/52 (2%) of patients with benign breast disease, 8/40 (20%) of all breast cancer patients, 6/19 (32%) of breast cancer patients with axillary nodal involvement and 1/1 of breast cancer patients with distant metastases were above this level. Among the same patients the CEA serum test was positive at a cut-off level of 5 micrograms/l in 7/40 (18%) cancer cases, and in 6/19 (32%) of cancer patients with nodal involvement. When we used the cut-off level 35 kU/l for CA 15-3 and 5 micrograms/l for CEA 1/52 (2%) of patients with benign breast disease, 10/40 (25%) of all breast cancer patients, 7/19 (37%) patients with axillary nodal involvement and 1/1 of breast cancer patients with distant metastases were positive in one or both of the tests. The serum CA 15-3 and CEA values were higher in patients with tumour size above 2 cm in diameter than in patients with smaller tumours (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

A new tumor marker MCA in breast cancer diagnosis.

The serum concentration of the new tumor marker MCA (mucinous carcinoma associated antigen) was determined by an enzyme immunoassay kit method, which is based on the use of a monoclonal antibody b12. The mean MCA values in breast cancer patients (n = 40) were significantly higher than in patients with benign breast disease (n = 55, p less than 0.001) and in control subjects (n = 37, p less than 0.001). When we used the cut-off level 11 KU/l for MCA, 6/37 (16.2%) of control subjects 7/55 (12.7%) of patients with benign breast disease, 18/40 (45.0%) of all breast cancer patients 11/19 (57.9%) of breast cancer patients with axillary node involvement, and 1/1 breast cancer patient with distant metastases were above this cut-off level. For comparison, at the cut-off level of 5 micrograms/l the CEA test was positive in 7/40 (18%) cancer cases, and in 6/19 (32%) of cancer patients with nodal involvement. Patients with axillary nodal metastasis showed higher values than patients without metastasis in both tests (p less than 0.01). The combination of MCA at 11 KU/l cut-off level and CEA at the 5.0 micrograms/l cut-off level reached the diagnostic sensitivity of 0.53, efficiency of 0.73, and specificity of 0.87. It seems that MCA is a promising tumor marker in breast cancer. Especially high values may have diagnostic significance.

Performance of direct equilibrium dialysis and analogue-type free thyroid hormone assays, and an immunoradiometric TSH method in patients with thyroid dysfunction.

Direct equilibrium dialysis and analogue-type radio-immunoassays for free triiodothyronine (FT3) and free thyroxine (FT4) in serum were compared in 168 subjects with various states of thyroid function. A good diagnostic efficacy for FT3 and FT4 by either type of assay was observed in hyperthyroidism. In hypothyroidism the free thyroid hormone assays, particularly the FT3 assays, performed diagnostically less well, partly because patients with mild disease were included in the study. No significant differences in the percentages of misclassifications of thyroid dysfunction patients by corresponding dialysis and analogue assays were found. We observed a good linear correlation between dialysis and analogue methods for FT3 (r = 0.98) and FT4 (r = 0.97) in this study comprising out-patients not suffering from severe non-thyroidal disease, known from earlier studies in this and other laboratories to interfere in these assays. It is concluded that analogue assays may be used on out-patients in whom severe systemic diseases are less frequent than in hospitalized patients. There are, however, other limitations to the use of analogue assays than systemic diseases. We observed two euthyroid patients with thyroxine auto-antibodies causing very high FT4 concentrations as determined by analogue assay; their dialysable FT4 concentrations were normal. We also tested a recently developed immunoradiometric serum TSH assay, which was found to perform well in primary hypo- and hyperthyroidism. Serum TSH was elevated in one patient hyperthyroid because of a TSH-producing pituitary adenoma, and within the reference limits in a patient with secondary hypothyroidism.

Concentrations of iodothyronines in serum of patients with chronic renal failure and other nonthyroidal illnesses: role of free fatty acids.

The mean concentration of free thyroxin (FT4) in serum, as determined by direct equilibrium dialysis, was decreased in patients with chronic renal failure (CRF) and increased in patients with various other nonthyroidal illnesses (NTI). The mean concentration of dialyzable free triiodothyronine (FT3) in serum was equally low in both groups of patients. Patients with CRF of various etiology but a similar degree of renal failure as estimated from serum creatinine assay had very similar concentrations of FT4 and FT3 in their serum. Mean thyroxin (T4) and triiodothyronine (T3) concentrations in serum were decreased in CRF and NTI, whereas the mean reverse-T3 concentration in serum was normal in CRF and increased in NTI. T4-binding globulin and albumin were markedly decreased in CRF and NTI; T4-binding prealbumin was increased in CRF and decreased in NTI. The mean concentration of nonesterified free fatty acids (FFA) in serum was increased in NTI but not in CRF. The weak, but significant, positive correlation observed between FT4 and FFA in serum (r = 0.34, P less than 0.01) in NTI indicates that the increase in serum FT4 in this group of patients could be an effect, at least in part, of FFA competing with T4 for binding sites on serum proteins. The stronger correlation detected between the serum FT4 concentration and the FFA/albumin molar ratio in serum (r = 0.60, P less than 0.001) demonstrates the importance of a low albumin concentration for expression of the effect of FFA on FT4 in severe systemic illnesses.

Cerebrospinal fluid lysozyme and beta 2-microglobulin in neurosarcoidosis.

The levels of lysozyme (LZM) and beta 2-microglobulin (beta 2m) were measured in the cerebrospinal fluid (CSF) and serum of 32 patients with sarcoidosis, 20 of whom had neurosarcoidosis. LZM was analyzed by a new radioimmunoassay (RIA) modification. CSF LZM was elevated in 15 of 20 patients with neurosarcoidosis but in only 4 of 12 patients with extraneural sarcoidosis. CSF beta 2m values were elevated in 13 of 19 and in one of 11 patients, respectively. In neurosarcoidosis, both CSF LZM and beta 2m correlated to CSF leucocytes but not significantly to CSF albumin thus suggesting that LZM and beta 2m were secreted from cells within the central nervous system (CNS). In patients with sarcoidosis, elevations of CSF LZM and beta 2m revealed disease activity in the CNS. Both analyses were also useful in the follow-up of neurosarcoidosis.

A sensitive and practical immunoradiometric assay of thyrotropin.

We describe a two-site immunoradiometric assay for thyrotropin (TSH) in serum, based on use of two monoclonal antibodies directed against two separate antigenic determinants on the TSH molecule. One antibody is immobilized on polystyrene beads; the other is radioiodinated by a modified Chloramine T method. The detection limit of the assay is 0.02 milli-int. unit/L. The working range (CV less than 10%) is from 0.1 to greater than 50 milli-int. units/L. The log mean concentration of TSH in sera collected from 100 euthyroid subjects between 08:00 and 11:00 hours was 1.9 milli-int. units/L, the range 0.4-5.4 milli-int. units/L. Values for hyperthyroid patients and thyroid-cancer patients being treated with thyroxin were much lower than those for euthyroid persons. Results by this new assay correlated excellently with those by our conventional radioimmunoassay (r = 0.99) and also with a sensitive immunofluorometric TSH method (Delfia TSH) (r = 0.99).