Tear fluid concentration of mmp–8 is elevated in non—allergic eosinophilic conjunctivitis and correlates with conjunctival inflammatory cell infiltration

Background  To investigate tear fluid concentration of matrix metalloproteinase 8 (MMP–8) and its relation to conjunctival inflammatory cell infiltration in persistent non—allergic eosinophilic conjunctivitis (NAEC). Methods  Two groups were included: 26 consecutive adult patients with NAEC (conjunctival eosinophils at least 1+ [1-10 eosinophils/slide], skin prick test [SPT] to common allergens negative), and 26 asymptomatic adult persons (no conjunctival eosinophils, SPT negative). MMP–8 tear fluid concentrations were determined by immunofluorometric assay, and conjunctival brush cytology samples from NAEC patients were used for MMP–8 immunocytochemistry. Gelatin zymography was used to illustrate proteolytic activity within the tear fluid samples. Results  The mean MMP–8 concentration was significantly higher among NAEC patients (214.3 ± 327.7 μg/l) than among healthy persons (50.4 ± 62.3 μg/l, P < 0.0001). In the NAEC patients, tear fluid MMP–8 correlated with the numbers of conjunctival neutrophils (r = 0.66, P = 0.0002) as well as with goblet cells and columnar epithelial cells (r = 0.54 for both, P = 0.045), but not with the lymphocyte numbers (r = -0.36, P = 0.0741). By immunocytology, MMP–8 protein could also be detected in vivo in the inflammatory cell population within the conjunctiva. Zymography revealed that proteolysis was significantly higher in the NAEC group, and activated enzymes were practically found only in the NAEC group. Conclusions  The results showed that NAEC is an inflammatory condition characterized by increased tear fluid MMP–8 levels, probably derived from both inflammatory and structural conjunctival cells. The increased proteolytic activity in NAEC patients may indicate risk of conjunctival structural changes (remodeling).     

Matrix metalloproteinase-8 (MMP-8) is the major collagenase in human dentin

Objective

Previously an unidentified collagenolytic metalloprotease together with gelatinase (matrix metalloproteinase-2, MMP-2), and enamelysin (MMP-20) have been detected in human dentin. The aim of the study was to characterize dentinal collagenolytic enzymes. Furthermore, we hypothesized that the dentinal MMPs are protected by the mineral phase, and studied the stability of dentinal MMPs.

Design

To characterize dentinal collagenolytic enzymes, we used Western blotting with specific antibodies against MMP collagenases (MMP-1, -8, and -13) and cathepsin K. MMP-8 immunofluorometric assay (IFMA) was also used for MMP-8 detection, and functional collagenase activity was examined with type I collagen degradation assay. The stability of dentinal MMPs was examined by autoclaving dentin blocks before protein extraction and subsequent examination of protein levels and the activities of dentin collagenase and gelatinases.

Results

MMP-8 (collagenase-2) was detected in dentin both with Western blot and IFMA, and dentinal samples also cleaved the intact type I collagen into characteristic 3/4(αA)-cleavage products in vitro. No other collagenases or cathepsin K were detected. In autoclaved samples no MMP-8 was found, but gelatinase activity was observed in protein fractions of mineralized dentin.

Conclusions

MMP-8 represents the major collagenase in human dentin. Unlike MMP-8, dentinal gelatinases can be detected after autoclave treatment of dentin, indicating their high resistance to external sample treatment procedures.     

Wide-band spectral analysis of blood pressure and RR interval variability in borderline and mild hypertension.

The aim of this study was firstly to investigate whether indices of wide-band spectral analysis in borderline hypertensive (BHT) or mildly hypertensive (HT) subjects differ from those in normotensive (NT) subjects, and secondly to assess the predictive value of these indices for future hypertension. Electrocardiogram and intra-arterial 24 h ambulatory blood pressure (BP) were recorded in 32 NT, 29 BHT and 30 HT middle-aged men. From the recordings, a 16 h period was extracted for wide-band spectral analysis. A single spectrum of BP and RR interval (RRI) variability was computed for each period by the fast Fourier transform method. The slopes of the spectra were assessed on a log-log scale by linear fitting of the spectral values. Power spectral densities were calculated over regions of 0-0.003, 0.003-0.04, 0.04-0.15, 0.15-0.40 and 0-0.4 Hz. No between-group differences were found in the slopes of BP and RRI spectra. The between-group differences in spectral powers for BP variability were almost invariably significant. The spectral powers for RRI variability did not show between-group differences. Five years later, 22 NT, 22 BHT and 18 HT subjects were re-assessed using casual BP measurements. In a logistic regression model for the combined group of NT and BHT subjects who became HT (22 of 44) during the five-year period, none of the parameters of wide-band spectrum predicted the development of hypertension. In conclusion, parameters of wide-band spectral analysis may not be useful in predicting future hypertension in NT and BHT subjects. Because the BP level is a major factor influencing BP variability, the between-group differences in wide-band spectral powers in BP may be due to differences in BP level rather than differences in cardiovascular regulatory mechanisms.     

Expression of pertussis toxin subunit S4 as an intracytoplasmic protein in Bacillus subtilis

The expression and secretion of pertussis toxin subunits S1 to S5 in Bacillus subtilis by the aid of a bacillar signal sequence has been reported. While secretion of subunit S1 was high, that of others was low. Ways have now been explored to improve the yield, using S4 as an example. The addition of a protease inhibitor was found to increase the amount of S4 in the culture supernatant, but the final amount was still much below that of S1. However, intracellular expression of S4 gave a high yield (500 mg I⁻¹) and the aggregated protein could easily be isolated in a few simple steps.     

Macrophage-colony stimulating factor (M-CSF) is increased in the synovial-like membrane of the periprosthetic tissues in the aseptic loosening of total hip replacement (THR)

Summary The aim of the study was to assess the eventual presence, cellular localization and extent of expression of the osteoclast activating cytokine M-CSF (CSF-1) in the periprosthetic tissues around loose total hip replacement (THR). Synovial-like membrane was obtained from the implant-to-bone interface and pseudocapsule from ten total hip revisions performed for aseptic loosening and compared to ten hip synovial tissue samples obtained from ten patients who had primary THR for osteoarthritis. Avidin-biotinperoxidase complex (ABC) and alkaline phosphatase-anti-alkaline phosphatase (APAAP) methods were used for staining and VIDAS image analysis for quantification. M-CSF was mainly produced by macrophages, which often contained wear particles, but also by some fibroblasts and vascular endothelial cells. The number of cells containing (per one mm² tissue) clearly increased in the interface (1585±212; p<0.01) and pseudocapsular (1456±248; p<0.01) tissue compared to synovial tissue (543±118). The present findings suggest, that inflammatory foreign-body type of response enhances expression of M-CSF in cases of aseptic loosening of THR. M-CSF produced in the synovial-like membrane in the implant-bone interface may contribute to activation of osteoclasts in periprosthetic bone and thus to loosening.     

Collagenase 2/matrix metalloproteinase 8 in critically ill patients with secondary peritonitis

Secondary peritonitis is an important indication for surgical intensive care admissions, and it is associated with high morbidity and mortality. Collagenase 2/matrix metalloproteinase (MMP) 8 is a tissue matrix-degrading enzyme that is released from leukocytes upon inflammatory stimuli and may thus contribute to peritonitis-associated organ damage. We studied the levels and activity of MMP-8 in the peritoneal fluid of 15 critically ill patients with secondary peritonitis. The MMP-8 levels were measured from the patients' peritoneal fluid, serum, and urine, and from the serum and urine of 10 healthy controls by immunofluorometric assay. Median MMP-8 level in peritoneal fluid supernatant was 1,317 microg/L (interquartile range [IQR]) (1,254-1,359 microg/L) being significantly higher than in the sera of the patients (P=0.008). Molecular forms and isoform distribution of MMP-8, MMP-1, and MMP-13 in peritoneal fluid, assessed by Western immunoblotting, revealed that the neutrophil-type MMP-8 was the major collagenase species in peritoneal fluid, and it was partially in an activated form. Catalytically competent, active MMP-8 produced the characteristic cleavage products from intact human type I collagen. The serum levels of MMP-8 were higher in the patients, 49 microg/L (IQR, 23-214 microg/L), than in the controls, 11 microg/L (IQR, 8-24 microg/L) (P<0.01). The MMP-8 levels in the urine were higher in the patients, 0.27 microg/L (IQR, 0.04-1.89 microg/L), than in the controls, 0.03 microg/L (IQR, 0.0-0.05 microg/L) (P=0.013). Our data demonstrate for the first time that MMP-8 levels are remarkably elevated and in an active and catalytically competent form in the peritoneal fluid samples of patients with secondary peritonitis.     

Association of the salivary triggering receptor expressed on myeloid cells/its ligand peptidoglycan recognition protein 1 axis with oral inflammation in kidney disease

1 Background

Triggering receptor expressed on myeloid cells (TREM‐1) is a cell‐surface receptor involved in amplification of inflammatory response to bacterial infections, along with its ligand peptidoglycan recognition protein 1 (PGLYRP1). TREM‐1 is shed by matrix metalloproteinases (MMPs) to its soluble (s) form. The aim of the study is to investigate association of sTREM‐1 and PGLYRP1 with oral inflammatory burden among patients with chronic kidney disease (CKD) at predialysis and posttransplantation stages.

2 Methods

One hundred forty‐four patients with CKD were examined at predialysis, and oral infection foci were treated prior to kidney transplantation. Fifty‐three patients were available for follow‐up after transplantation. Oral inflammatory burden was assessed by the Periodontal Inflammatory Burden Index (PIBI) and Total Dental Index. sTREM‐1, PGLYRP1, and interleukin (IL)‐1β were measured in saliva by enzyme‐linked immunosorbent assay, and MMP‐8 was measured by immunofluorometric assay.

3 Results

In the predialysis stage, sTREM‐1 and PGLYRP1 were positively associated with IL‐1β, MMP‐8, and PIBI. More specifically, patients with deeper probing depth (PD) (at least two sites with ≥6 mm) had higher concentrations of salivary sTREM‐1 and PGLYRP1 compared with those with shallower PD. Higher concentrations of PGLYRP1 and IL‐1β were associated with a higher number of teeth (> 25). On follow‐up, higher PGLYRP1 and sTREM‐1 were associated with one or more sites with ≥4 mm PD.

4 Conclusions

sTREM‐1 and PGLYRP1 are elevated in patients with CKD with poor oral health and positively correlate with number of active periodontal pockets after oral infection therapy. Moreover, they positively correlate with MMP‐8 and IL‐1β. Hence, the salivary sTREM‐1/PGLYRP1 axis could be useful as a diagnostic marker for oral infection within patients with CKD.     

Otitis externa sicca/fibrotising external otitis (FEO) as a complication of Sjögren's syndrome

Sj?gren's syndrome (SS) is a condition characterized by sicca symptoms and by autoimmune features. We describe two SS patients with otitis externa fibroticans/sicca. One of these 2 patients developed a lesion of the tympanic membrane making it necessary to perform a tympantomy and meatoplasty. Our findings suggest firstly that the epithelial cell-mediated secretion of lamellar bodies and the production of the permeability barrier are defective in SS. Secondly, local moisturing and/or topical corticosteroid treatment in SS patients with sicca symptoms in the auditory canal could help to avoid reconstructive surgical treatment.