PURPOSE: The purpose of this study was to investigate the expression of cytokeratin (CK) 8 in the corneoconjunctival epithelium. METHODS: In 17 cadaveric corneoscleral discs and 3 other discs, the presence of CK8 alone or CK8, together with CK3, CK15, vimentin, and integrin α6, was investigated by using indirect immunohistochemistry on radial cryosections. Four corneoscleral discs stored in organ culture were used for the preparation of tangential sections of the limbus and for the isolation of limbal epithelial cells and their subsequent cultivation. CK8 expression was examined by RT-PCR in the corneal, limbal, and conjunctival epithelium. RESULTS: Sixty percent of the cadaveric corneoscleral samples and all samples stored in organ culture revealed positivity for CK8 in the basal epithelial layer of the limbus. Positive basal cells formed a single line or separated clusters. The signal for CK8 became weaker toward the surface of the limbal epithelium. The colocalization of CK8 with vimentin and CK15 in the limbus was also found. CK3 showed only occasional positivity in some of the surface limbal cells. The expression of integrin α6 in the basal membrane was absent or decreased under the CK8-positive clusters. Cell cultures revealed strong positivity for CK8 in approximately 80% of the cultured cells, and CK8 expression in the cornea, limbus, and conjunctiva was determined by RT-PCR. CONCLUSIONS: The study demonstrates the strong expression of CK8 in limbal epithelial basal cells, which is maintained during the differentiation and migration of the limbal cells toward the central corneal epithelium.

• MeSH
• buněčné kultury MeSH
• dospělí MeSH
• epitelové buňky metabolismus   MeSH
• exprese genu MeSH
• fluorescenční mikroskopie MeSH
• fluorescenční protilátková technika nepřímá MeSH
• integrin alfa6 metabolismus   MeSH
• keratin-15 metabolismus   MeSH
• keratin-3 metabolismus   MeSH
• keratin-8 genetika   metabolismus   MeSH
• konjunktiva cytologie   metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• limbus corneae cytologie   metabolismus   MeSH
• messenger RNA genetika   MeSH
• mladiství MeSH
• orgánové kultury - kultivační techniky MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• rohovkový epitel cytologie   metabolismus   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• vimentin metabolismus   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

PURPOSE: To investigate the contribution of matrix metalloproteinases (MMPs) to recurrent corneal melting in keratoconjunctivitis sicca associated with primary Sjorgen's syndrome (pSS). METHODS: One native melted cornea and ten melted corneal grafts from two patients with severe pSS were used. The presence of MMPs (1, 2, 3, 7, 8, 9, and 13) was detected using indirect enzyme immunohistochemistry. The active forms of MMP 2 and 9 and MMP 3 and 7 were examined by gelatin and casein zymography, respectively. The concentrations of active MMP 1 were measured using an activity assay. Eleven unaffected corneas served as controls. RESULTS: The average values of the staining intensity revealed very intense MMP 1, intense MMP 2, 7, and 9 and moderate MMP 3 and 8 positivity, in the corneal epithelium of melted corneas. Intense MMP 1 and 9 staining, moderate MMP 2, 3, and 8 staining, and weak MMP 7 staining were found in the anterior stroma. The posterior stroma revealed intense MMP 1, moderate MMP 3 and 9, and weak MMP 2, 7, and 8 positivity. Immunostaining of the endothelium was moderate for MMP 9 and weak for MMP 1, 2, 3, 7, and 8. MMP 13 was negative in all but four melted specimens, where weak-to-moderate staining was found in the epithelium and stroma. Control corneas revealed moderate MMP 1 and 2 and weak MMP 8 staining in the epithelium, weak MMP 2 staining in the anterior stroma, and weak MMP 1 and 8 staining in the endothelium. Significantly elevated MMP 1 activity and extremely elevated MMP 9 activity were found in most of the tested pathological specimens, compared to healthy controls, where no activity of the two enzymes was present. Markedly elevated MMP 2 activity was found in 2 of 11 specimens, compared to normal tissue. The inactive form of MMP 3 was detected in half of the tested specimens, and inactive MMP 7 in all melted corneas. Active MMP 3 and 7 were found in one melted sample. Neither of these MMPs was found in any of the control specimens. CONCLUSIONS: The increased expression and elevated activity of a wide range of MMPs in melted cornea samples from two patients diagnosed with pSS confirm that these enzymes contribute to the tissue destruction, leading to serious consequences such as corneal perforation and loss of vision.

• MeSH
• imunohistochemie MeSH
• lidé středního věku MeSH
• lidé MeSH
• matrixové metaloproteinasy metabolismus   MeSH
• nemoci rohovky enzymologie   patologie   MeSH
• rohovka enzymologie   patologie   MeSH
• senioři MeSH
• syndrom MeSH
• transport proteinů MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• kazuistiky MeSH
• práce podpořená grantem MeSH