SPINT2 (HAI-2) missense variants identified in congenital sodium diarrhea/tufting enteropathy affect the ability of HAI-2 to inhibit prostasin but not matriptase.

The syndromic form of congenital sodium diarrhea (SCSD) is caused by bi-allelic mutations in SPINT2, which encodes a Kunitz-type serine protease inhibitor (HAI-2). We report three novel SCSD patients, two novel SPINT2 mutations and review published cases. The most common findings in SCSD patients were choanal atresia (20/34) and keratitis of infantile onset (26/34). Characteristic epithelial tufts on intestinal histology were reported in 13/34 patients. Of 13 different SPINT2 variants identified in SCSD, 4 are missense variants and localize to the second Kunitz domain (KD2) of HAI-2. HAI-2 has been implicated in the regulation of the activities of several serine proteases including prostasin and matriptase, which are both important for epithelial barrier formation. No patient with bi-allelic stop mutations was identified, suggesting that at least one SPINT2 allele encoding a protein with residual HAI-2 function is necessary for survival. We show that the SCSD-associated HAI-2 variants p.Phe161Val, p.Tyr163Cys and p.Gly168Ser all display decreased ability to inhibit prostasin-catalyzed cleavage. However, the SCSD-associated HAI-2 variants inhibited matriptase as efficiently as the wild-type HAI-2. Homology modeling indicated limited solvent exposure of the mutated amino acids, suggesting that they induce misfolding of KD2. This suggests that prostasin needs to engage with an exosite motif located on KD2 in addition to the binding loop (Cys47/Arg48) located on the first Kunitz domain in order to inhibit prostasin. In conclusion our data suggests that SCSD is caused by lack of inhibition of prostasin or a similar protease in the secretory pathway or on the plasma membrane.

Insights into the regulation of the matriptase-prostasin proteolytic system.

The membrane-associated prostasin and matriptase belonging to the S1A subfamily of serine proteases, are critical for epithelial development and maintenance. The two proteases are involved in the activation of each other and are both regulated by the protease inhibitors, HAI-1 and HAI-2. The S1A subfamily of serine proteases are generally produced as inactive zymogens requiring a cleavage event to obtain activity. However, contrary to the common case, the zymogen form of matriptase exhibits proteolytic activity, which can be inhibited by HAI-1 and HAI-2, as for the activated counterpart. We provide strong evidence that also prostasin exhibits proteolytic activity in its zymogen form. Furthermore, we show that the activity of zymogen prostasin can be inhibited by HAI-1 and HAI-2. We report that zymogen prostasin is capable of activating zymogen matriptase, but unable to activate its own zymogen form. We propose the existence of an unusual enzyme-enzyme relationship consisting of proteolytically active zymogen forms of both matriptase and prostasin, kept under control by HAI-1 and HAI-2, and located at the pinnacle of an important proteolytic pathway in epithelia. Perturbed balance in this proteolytic system is likely to cause rapid and efficient activation of matriptase by the dual action of zymogen matriptase and zymogen prostasin. Previous studies suggest that the zymogen form of matriptase performs the normal proteolytic functions of the protease, whereas excess matriptase activation likely causes carcinogenesis. HAI-1 and HAI-2 are thus important for the prevention of matriptase activation whether catalysed by zymogen/activated prostasin (this study) or zymogen/activated matriptase (previous studies).

Inhibition of an active zymogen protease: the zymogen form of matriptase is regulated by HAI-1 and HAI-2.

The membrane-bound serine protease matriptase belongs to a rare subset of serine proteases that display significant activity in the zymogen form. Matriptase is critically involved in epithelial differentiation and homeostasis, and insufficient regulation of its proteolytic activity directly causes onset and development of malignant cancer. There is strong evidence that the zymogen activity of matriptase is sufficient for its biological function(s). Activated matriptase is inhibited by the two Kunitz-type inhibitor domain-containing hepatocyte growth factor activator inhibitors 1 (HAI-1) and HAI-2, however, it remains unknown whether the activity of the matriptase zymogen is regulated. Using both purified proteins and a cell-based assay, we show that the catalytic activity of the matriptase zymogen towards a peptide-based substrate as well as the natural protein substrates, pro-HGF and pro-prostasin, can be inhibited by HAI-1 and HAI-2. Inhibition of zymogen matriptase by HAI-1 and HAI-2 appears similar to inhibition of activated matriptase and occurs at comparable inhibitor concentrations. This indicates that HAI-1 and HAI-2 interact with the active sites of zymogen and activated matriptase in a similar manner. Our results suggest that HAI-1 and HAI-2 regulate matriptase zymogen activity and thus may act as regulators of matriptase trans(auto)-activation. Due to the main localisation of HAI-2 in the ER and HAI-1 in the secretory pathway and on the cell surface, this regulation likely occurs both in the secretory pathway and on the plasma membrane. Regulation of an active zymogen form of a protease is a novel finding.

The Rotoglide™ total replacement of the first metatarso-phalangeal joint. A prospective series with 7-15 years clinico-radiological follow-up with survival analysis.

BACKGROUND: The Rotoglide total replacement of the MTP-1 joint. 15 years survival analysis. The purpose of this prospective study was to evaluate the long-term performance clinico-radiographically of an uncemented three-component total replacement for the first metatarso-phalangeal joint (MTP-1) used for hallux rigidus (primary osteoarthritis grades 3 and 4). The follow-up was median 11.5 years (7-15). METHODS: The AOFAS forefoot score was used preoperatively and at follow-up. Radiographs were taken weight-bearing in the AP-projection and in tip-toe standing in the lateral view. Arthrosis in the sesamoid junction, prosthetic loosening, subsidence (of prosthesis as well as sesamoids), and dorsiflexion were measured, recorded and subjected to multiple variance analysis. Survival analysis was performed for 15 years. MATERIAL: Ninety implants in 80 patients (53 women and 27 men); median age 58 (41-76) were evaluated. RESULTS: Six patients representing seven prostheses in situ had died from unrelated reason. The median preoperative AOFAS increased significantly from 40 to 95. The median gain was 45. Four replacements (4.4%) were extracted for other reasons than loosening. No aseptic loosenings were recorded. The survival rate at 15 years was 91.5% (83-100). Multiple variance analysis showed that arthrosis in the metatarso-sesamoid junction correlated with reduced AOFAS score. CONCLUSION: The prosthesis has stood the test of time; the results justify its further use.

Conservatively treated wound for four months turned out to be highly malignant cutaneous angiosarcoma.

Angiosarcomas are rare, highly malignant tumours of vascular origin. They present as fast growing, haematoma-like and diffuse lesions. In this case report, a 71-year-old woman presented with what was assessed as an abscess arising from a traumatic haematoma of the forehead. The lesion was primarily treated conservatively. The condition progressed despite of treatment to a point where the tumour covered a third of her face. After four months, skin biopsies were taken and the diagnosis angiosarcoma was made. This case report should lead to an increased focus on angiosarcomas and the importance of sufficient biopsies.

[Basic registration of risk factors in medical records. Malnutrition, overweight, physical inactivity, smoking and alcohol]. / Basale journaloplysninger om risikofaktorer. Underernaering, overvaegt, fysisk inaktivitet, rygning og alkohol.

INTRODUCTION: Lifestyle factors are important for clinical outcome. Systematic and early identification of these factors is important in order to offer relevant lifestyle intervention programmes. The objective was to evaluate whether basic registration of risk factors was understandable, applicable and sufficient in the clinical workday. MATERIALS AND METHODS: Eleven clinical specialists participated from a broad range of departments. They performed identification and registration of malnutrition, overweight, physical inactivity, smoking and harmful alcohol consumption based on medical records from own departments (in total 2420 times). The specialists then evaluated the understanding, applicability and sufficiency (363 times). Their comments were noted. RESULTS: Identification and registration was accomplished for 85% (0-100%) of the records. Except for two fields--"measurements of waist" and "other training programs"--the specialists found the basic registration understandable, applicable and sufficient. They lacked more details regarding stress-response in the information material and pointed out inconsistencies in the wording. Ten specialists found that the recommended clinical approach was possible to implement in the present routines. CONCLUSION: Basic registration of lifestyle factors for adult patients is possible and clinical relevant. It is recommended that the Danish National Board of Health uses the results in future.