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Biomedical subjects

Timo Tervo

Publications and source records attributed to Timo Tervo.

18 recordsLinked to original sources
PubMed2006

Cooperation of isoforms of laminin-332 and tenascin-CL during early adhesion and spreading of immortalized human corneal epithelial cells.

The repair of corneal wounds requires both epithelial cell adhesion and migration. We have studied the early adhesion process of immortalized human corneal epithelial (HCE) cells and show by field emission scanning electron microscopy (FESEM) that the cells first adhere via foot-like process to the growth substratum and later present lamellar spreading. During early adhesion indirect immunofluorescence showed that the cells codeposited laminin (Lm) -332 and the large subunit of tenascin-C (Tn-CL) as a demarcated plaque beneath the cells. Instead, unprocessed Lm-332 (alpha 3'32) was found in a wider area in cells showing lamellar spreading and was also prominently expressed in the cytoplasm of the migrating marginal cells in the in vitro wounded HCE cultures. Confocal laser scanning microscopy (CLSM) showed that the Golgi apparatus was located to the vicinity of the Lm-332/Tn-CL-containing adhesion plaque and accordingly treatment of the cells with demecolcine, dispersing the Golgi apparatus, prevented the formation of plaques. This suggests that formation of the adhesion plaque depends on a direct vectorial secretion of Lm-332 and Tn-CL to the culture substratum. Instead, cytochalasin B treatment disrupted microfilaments and arborized the cells but did not affect the deposition of Tn-CL/Lm-332 as a plaque beneath the cells. The suggestion was supported by immunoprecipitation experiments which showed that Tn-CL and Lm alpha 3' chain were found in cell-free matrices on the culture substratum of spreading cells but not at all (Tn-CL) or much less (Lm-332) in the culture medium. Quantitative cell adhesion experiments showed that HCE cells did not adhere to plain Tn-C coat and that integrin (Int) alpha(3)beta(1) mediated the adhesion of HCE cells to purified Lm-332 and to Lm-332/Tn-C while Int beta4 did not mediate adhesion to these proteins. Taken together, our data suggest that Lm-332 and Tn-CL cooperate in early adhesion process of HCE cells. Furthermore, the results show that Lm-3'32 isoform functions in the spreading of the cells beyond the early adhesion stage and appears to emerge into HCE cells starting to migrate in experimental wounds.

Cell Adhesion↗
PubMed2006

Differential expression of collagen types XVIII/endostatin and XV in normal, keratoconus, and scarred human corneas.

PURPOSE: This study was designed to clarify the expression of 2 closely related collagen (Col) types XVIII and XV, and the proteolytically derived endostatin fragment of ColXVIII in normal, keratoconus, and scarred human corneas. METHODS: Immunohistochemistry, in situ hybridization, immunoelectron microscopy, and Western immunoblotting were used for human corneal samples obtained from penetrating keratoplasty. RESULTS: In the normal cornea, ColXVIII was immunolocalized to the corneal and conjunctival epithelial basement membrane (EBM), Descemet s membrane, and the limbal and conjunctival capillaries. Immunoreaction for endostatin was otherwise similar, but it also was present in corneal epithelial cells. Western immunoblotting showed that normal cornea contains several endostatin fragments ranging from 20 to 100 kDa. ColXV was present in the EBM of the limbus and conjunctiva, but not in EBM of the clear cornea. In situ hybridization revealed that corneal basal epithelial cells were responsible for the synthesis of ColXVIII mRNA. Keratoconus cases were characterized by an irregular EBM immunoreactivity for ColXVIII and endostatin and patchy immunoreactivity beneath EBM. In scarred corneas, highly increased immunoreactivity for ColXVIII, endostatin, and ColXV was present within stroma. CONCLUSIONS: The results indicate that ColXVIII and ColXV are differentially expressed in normal human corneas. Constant expression of ColXVIII by corneal EBM suggests that it is an important structural molecule. Aberrant expression of ColXVIII, endostatin, and ColXV in keratoconus and scarred corneas emphasizes the active role these molecules in the wound healing process.

Adult↗
PubMed2006

EMMPRIN and MMP-1 in keratoconus.

PURPOSE: This study was conducted to determine the eventual presence and activity of EMMPRIN (extracellular matrix metalloproteinase inducer CD147) and interstitial collagenase MMP-1 in the cornea of keratoconus patients and their eventual interrelationship. MMP-1 was chosen because it is able to degrade fibrillar corneal collagens type I and III and might therefore play a role in stromal thinning in keratoconus. METHODS: Immunohistochemical labeling of EMMPRIN and MMP-1 in relation to histopathological changes in 5 keratoconus patients and 5 matched healthy controls was investigated. RESULTS: Relatively strong EMMPRIN expression was found in normal corneal epithelial cells, but moderate expression was also found in stroma. In keratoconus, EMMPRIN was found in all layers of cornea, especially in histopathologically altered areas. In normal cornea, MMP-1 staining was weak and restricted to epithelial cells. In keratoconus, MMP-1 expression was slightly augmented in epithelial cells and also appeared locally in a scattered manner in the stroma. The distribution of MMP-1 did not totally overlap with that of histologically apparent corneal damage and EMMPRIN expression. CONCLUSIONS: Both EMMPRIN and MMP-1 are upregulated in keratoconus, but MMP-1 may not be the only tissue destructive MMP upregulated by EMMPRIN as only EMMPRIN expression correlated topologically very well with corneal damage.

Basigin↗
PubMed2006

Altered expression of type XIII collagen in keratoconus and scarred human cornea: Increased expression in scarred cornea is associated with myofibroblast transformation.

PURPOSE: Type XIII collagen (ColXIII) is a transmembrane protein thought to be involved in cell-cell and cell-matrix interactions. We report here on its presence in the normal human cornea and compare the results for keratoconus and scarred corneas. METHODS: Immunohistochemistry and in situ hybridization were applied to human corneal samples obtained by penetrating keratoplasty. RESULTS: In the normal human cornea, ColXIII was immunolocalized to the corneal epithelial cells, and to a lesser degree to the stromal keratocytes. The keratoconus cases showed otherwise similar results, but in areas containing Bowman membrane disruptions showed thinned epithelial cells reduced immunostaining for ColXIII, whereas occasionally pronounced immunoreactivity was seen in the stromal keratocytes. The corneal scar samples contained highly increased ColXIII immunostaining by stromal cells in the fibrotic foci, whereas the peripheral areas showed less intense immunostaining. In situ hybridization confirmed that the corneal epithelium and keratocytes actively synthesize the transcript. Immunostaining with alphaSMA revealed that a substantial proportion of the ColXIII mRNA-expressing cells in the stromal scar tissues was myofibroblasts and that these areas lack CD34 immunoreactivity. CONCLUSIONS: The results indicate that ColXIII, which is predominantly confined to the basal corneal cells in the normal cornea, may have a role in the adhesion of corneal epithelial cells to each other and to the underlying basement membrane. Additionally, highly increased expression in scarred corneas suggests that it participates in the corneal wound healing process.

Actins↗
PubMed2006

Collagenolytic proteinases in keratoconus.

PURPOSE: To study the proteolytic phenomena contributing to the pathogenesis of keratoconus, corneal enzymes with potential to cleave fibrillar collagen were studied. METHODS: Immunohistochemical labeling was undertaken of conventional and novel mammalian collagenases (MMP-1, -2, -8, -13, and -14) of the matrix metalloproteinase (MMP) family and other collagenolytic proteinases of the serine (human trypsin-2) and cysteine (cathepsin K) endoproteinase families. The results were analyzed using a semiquantitative scoring system. RESULTS: Labeling of MMP-8 was moderate in healthy controls, but weak in keratoconus. Moderate MMP-2 and weak MMP-14 expressions were similar in controls and keratoconus. MMP-1 was slightly overexpressed in keratoconus. In contrast, MMP-13 was weak in controls compared to moderate in keratoconus and human trypsin-2 and cathepsin K were moderate in controls and strong in keratoconus. CONCLUSIONS: The collagenolytic milieu of human cornea is more complex than expected. Mesenchymal isoform of MMP-8 (ie, collagenase-2) participates in normal tissue remodeling, which may be impaired in keratoconus. MMP-2 (gelatinase A with interstitial collagenase activity) and MMP-14 (membrane-type MMP type I with collagenolytic potential) seem to be constitutively expressed and probably play a role in normal corneal remodeling. The most prominent changes in keratoconic cornea were observed in collagenase MMP-13 (ie, collagenase-3), and particularly, in cathepsin K and human trypsin-2, which were strongly expressed in keratoconus suggesting a role in intra- and extracellular pathological collagen destruction, respectively. This may contribute to stromal thinning characteristic for keratoconus.

Cathepsin K↗
PubMed2005

The immortalized human corneal epithelial cells adhere to laminin-10 by using Lutheran glycoproteins and integrin alpha3beta1.

We studied the adhesion characteristics of immortalized human corneal epithelial (HCE) cells on purified human laminin (Ln)-10 (alpha5beta1gamma1). Immunofluorescence studies with monoclonal antibodies (MAb) revealed the presence of laminin alpha5 chain receptor Lutheran (Lu) in vivo on the basal cells of cornea and as punctate cell surface-confined reactivity on adhering and spread HCE cells, while alpha- and beta-dystroglycans could not be found. Northern blot analysis showed the presence of 4.0 and 2.8 kb Lu transcripts in HCE cells. The results showed that Ln-10 induced adhesion and spreading of HCE cells without formation of focal adhesions. Quantitative adhesion assay showed that Lu together with integrin (Int) alpha3beta1 independently mediated the adhesion of HCE cells to Ln-10.

Blotting, Northern↗
PubMed2005

Nordic research in ophthalmology.

Nordic ophthalmologists and vision scientists are active in many fields of eye research. This is most evident at the biannual Nordic Congress of Ophthalmology, most recently held in Malmö in June 2004. The authors here review some of the research in vision and ophthalmology presented at this meeting or published recently by Nordic scientists. This paper does not represent a comprehensive review of all Nordic research in the field, but attempts to give an overview of some of the activities underway in eye research in this part of the world.

Animals↗
PubMed2004

Direct in vivo monitoring of acute allergic reactions in human conjunctiva.

Immediate allergic reactions are initiated by allergen-induced, specific IgE-mediated mast cell degranulation and involve leukocyte recruitment into the inflamed site. We compared conjunctival signs, symptoms, and in vivo leukocyte rolling and extravasation into sites of inflammation in five patients allergic to birch pollen and in 10 nonallergic controls who received a challenge to birch allergen or histamine. Both the specific allergen in allergic patients and histamine, both in patients and in healthy controls, induced symptoms and signs of an immediate allergic reaction together with leukocyte rolling within the conjunctival blood vessels. However, only allergen, not histamine, caused leukocyte extravasation into the site of inflammation in the allergic patients. Allergen also increased expression of endothelial P-selectin in conjunctival vessels and slowed the rolling of leukocytes which is required for their extravasation from blood circulation into the target tissue. Finally, i.v. heparin strongly reduced the number of slowly rolling cells during allergen- or histamine-induced reactions and this can probably hinder the leukocyte extravasation after allergen exposure. These findings suggest that slow rolling is required for leukocyte extravasation in acute allergic reactions, and it can be inhibited by heparin in vivo in therapeutically relevant conditions.

Acute Disease↗
PubMed2004

Monitoring leukocyte traffic in vivo into human delayed-type hypersensitivity reaction.

Leukocyte traffic from blood to sites of inflammation has been elaborately studied in animal models and in vitro. However, to date, little understanding has accumulated on the process in humans in vivo. A noninvasive light confocal microscopy technique has enabled us to image leukocyte rolling, arrest and transmigrated cells in vivo in human tissues. In the current study, a delayed-type hypersensitivity (DTH) reaction was elicited in the lower lip of healthy, Bacille Calmette-Guerin (BCG)-vaccinated volunteers by injection of respective purified protein derivate (PPD), mimicking the classic cutaneous Mantoux reaction. Subjects were imaged with real-time confocal microscopy at baseline and 2, 4, 24 and 48 h after the injection. The number of rolling leukocytes did not increase significantly until at 48 h. Even then, rolling cells were seen in only a minority (23%) of blood vessels. The frequency of engaged blood vessels was, nevertheless, significantly greater than at baseline. As the inflammation generated with this challenge was mild, transmigrated leukocytes could be detected in the confocal microscopy only occasionally. Histology of biopsies taken immediately after imaging at 48 h showed a T cell-dominated leukocyte population at the site of the DTH reaction. We have thus developed a method to monitor noninvasively leukocyte traffic in vivo in human subjects during the classic inflammatory model of delayed-type hypersensitivity reaction.

BCG Vaccine↗
PubMed2003

Leukocyte rolling and extravasation in surgical inflammation after mechanical and laser-induced trauma in human patients.

Acute inflammations main characteristics are exudation of fluid and plasma proteins and emigration of leukocytes, predominantly neutrophils into inflamed tissue. We have previously demonstrated that leukocyte extravasation can be directly monitored in human patients in vivo after tunnel cataract operation, but at the same time we noticed that only certain manipulations trigger also leukocyte extravasation. To define the type of surgical manipulations triggering leukocyte extravasation, we tested a set of surgical manipulations affecting distinct anatomical components of the conjunctiva. This set included various combinations of epithelial, nerve, and/or blood vessel damage. Manipulations tested here were surgical conjunctival biopsy, low-energy Argon laser treatment (80 mJ) closing a venule temporarily, high-energy argon laser treatment (480 mJ) to occlude a venule firmly, and 193 nm eximer laser-based conjunctival phototherapeutic keratectomy (160 mJ). These manipulations were compared to previous findings on conjunctival inflammation following the standard cataract operation. In mechanical trauma models (cataract operation and conjunctival biopsy) clinical signs of inflammation, number of leukocytes rolling, and number of tissue emigrated leukocytes were notably higher compared to pre-operative levels on one day after the operation. No specific anatomical triggering component for inflammation, including epithelial, nerve, and/or blood vessel damage, was indentified, but rather the trauma mechanism itself appeared to be an essential factor. Surprisingly, in laser-induced traumas no increase in number of rolling cells, rolling velocity slowing, or elevation of tissue emigrated leukocytes took place compared to pre-operative levels. These findings suggest that laser-induced traumas differ fundamentally from the mechanical traumas at all levels of leukocyte extravasation cascade, and at least in our setup, inflammatory reaction can be avoided or triggered depending on surgical method used.

Adult↗
PubMed2003

Group IIA phospholipase A(2) content in tears of patients having photorefractive keratectomy.

PURPOSE: To study the effect of photorefractive keratectomy (PRK) on the concentration of group IIA phospholipase A(2) (GIIAPLA(2)) in tears. SETTING: Departments of Ophthalmology and Pathology, University of Turku, Turku, and Helsinki University Eye Hospital, Helsinki, Finland. METHODS: Tear samples were collected from 25 eyes of 23 patients (mean age 32.3 years +/- 8.6 [SD]) preoperatively and 2 and 7 days after PRK. The GIIAPLA(2) concentration in the tears was measured by time-resolved fluoroimmunoassay. RESULTS: The GIIAPLA(2) concentration was significantly lower and the tear fluid flow rate significantly higher 2 days after PRK than preoperatively. At 7 days, the GIIAPLA(2) concentration and the tear fluid flow-corrected excretion of GIIAPLA(2) were significantly higher than preoperatively and at 2 days. The tear flow rate was also significantly higher than preoperatively. CONCLUSIONS: The GIIAPLA(2) content in tears decreased 2 days after PRK due to dilution of the GIIAPLA(2) content during hypersecretion of reflex tears. Photorefractive keratectomy caused an increase in the tear flow rate, GIIAPLA(2) concentration, and tear fluid flow-corrected excretion of GIIAPLA(2) in tears 7 days after surgery, enhancing the protection of tears against bacterial infections.

Adult↗
PubMed2003

In vivo confocal microscopy for evaluation of wound healing following corneal refractive surgery.

Understanding of corneal wound healing plays an important role, not only in management of corneal infections, but especially in refractive surgery. A better control of wound healing mechanisms might improve the results of such resculpturing techniques and help to avoid complications arising from these procedures. While studies have been focused in different aspects of corneal wound healing, our knowledge has increased greatly during the last years. Many problems associated with corneal healing also contribute to clinical pathology following corneal surgery. Understanding of such conditions has been augmented by the continuously developing corneal imaging techniques. We have used in vivo confocal microscopy (IVCM) for assessing corneas subjected to refractive surgery as well as corneas with common complications resulting from such procedures. IVCM has become a powerful tool for examining corneal cells, nerves, inflammations and infections. It allows information to be acquired repeatedly and at subbiomicroscopic levels that earlier had been obtainable only by invasive microscopic methods. Pre-examining corneas preoperatively by IVCM in order to reveal diseases or conditions in which elective refractive surgical procedures should not be undertaken or to select the ideal operation technique may help to avoid complications in the future. Measurement of the thickness of corneal sublayers or estimation of the thickness of a laser in situ keratomileusis flap or wound bed are other applications in which confocal microscopy may be valuable. In this article we attempt to describe the in vivo confocal findings of common refractive procedures and their complications, and discuss their biology in light of the existing knowledge on wound healing phenomena.

Animals↗
PubMed2003

Myopia and natural lighting extremes: risk factors in Finnish army conscripts.

PURPOSE: To establish whether features of environmental lighting in far northern latitudes might be associated with prevalence of myopia. METHODS: Using both questionnaires and military medical examinations, this cross-sectional survey of Finnish conscripts assessed both light exposure and conventional risk factors for myopia. RESULTS: While myopia was not associated with the month of birth, there was a trend towards a higher prevalence of myopia among conscripts living above the Arctic Circle, consistent with the hypothesis that ambient lighting might influence refractive development. Other novel associations with myopia were decreased sunglasses use and brown iris colour. As indicated by other reports, myopia was found to be associated with family history, education and nearwork. CONCLUSION: Although constraints inherent in surveying this military population may have limited our ability to detect associations, the positive findings suggest that studying northern populations may prove useful in clarifying any potential role of the light/dark cycle in refractive development.

Adolescent↗
PubMed2003

Production of fibronectin and tenascin isoforms and their role in the adhesion of human immortalized corneal epithelial cells.

PURPOSE: To study the production and deposition of fibronectin (Fn) isoforms and tenascin-C (Tn-C) by immortalized human corneal epithelial (HCE) cells and their integrin-dependent adhesion characteristics. METHODS: Indirect immunofluorescence with isoform-specific monoclonal antibodies (mAbs) was used to study extracellular matrix (ECM) protein composition and their integrin receptors in HCE cells. The synthesis of proteins was studied by Western blot analysis and adhesion by quantitative adhesion assay. RESULTS: HCE cells deposited fibrillar matrix containing extradomain EDA-Fn and sparser deposits of Onc-Fn, whereas Tn-C was deposited diffusely. EDA-Fn was present both in ECM and in culture medium, whereas Tn-C was present only in ECM. Fn-binding integrin (Int) alpha(5) subunit was present in subconfluent cells in focal adhesions (FAs) and matrix adhesions, whereas Int alpha(v)beta(5) was present in FAs in sparse cultures and as ringlike structures in denser cultures. Int alpha(v)beta(6) was colocalized with Int alpha(5) in FAs, only in cells adhering to growth substratum coated with Fn or Fn/Tn-C. Ints alpha(5)beta(1) and alpha(v)beta(6) mediated adhesion to Fn and Int alpha(v)beta(5) mediated adhesion to Vn, and both were inhibited by RGD peptide. The cells failed to adhere to Tn-C but adhered to Fn/Tn-C and were then more efficiently inhibited by the function-blocking integrin mAbs and RGD peptide. CONCLUSIONS: The results suggest corneal epithelial cells as the possible source for Fn isoforms and Tn-C in wound healing and pathologic conditions. The presence of Tn-C only in ECM suggest a vectorial deposition and adhesion experiments also indicate a role for Tn-C in Fn functioning.

Antibodies, Monoclonal↗
PubMed2002

A locus for autosomal dominant keratoconus: linkage to 16q22.3-q23.1 in Finnish families.

PURPOSE: The estimated world-wide prevalence of keratoconus is 50 to 230 per 100,000 in the general population. Sporadic keratoconus is the leading cause of corneal transplantation surgery in Western countries. Positive family history has been reported in 6% to 8% of patients. The purpose of this study was to map the disease locus in 20 Finnish families with autosomal dominant keratoconus, each family having two or more affected members and with no other associated genetic disease. METHODS: DNA was extracted from blood samples, collected from 42 affected and 34 unaffected family members. Genomic DNA from patients and their parents, was typed for alleles of 292 polymorphic markers. A genome-wide screening was performed to localize the disease gene. Fluorescent markers were amplified by polymerase chain reaction and separated on an automated sequencer. Allele sizes were assigned to each family member, after which LOD scores were calculated. RESULTS: The disease locus was mapped to chromosome 16q, between the markers D16S2624 and D16S3090, with a maximum parametric multipoint LOD score of 4.10 and corresponding nonparametric score of 3.27 (NPL, P = 0.00006). Evidence from 20 families provided support for the linkage, consistent with a single locus for familial autosomal dominant keratoconus without heterogeneity. CONCLUSIONS: This study is the first genome-wide linkage study to map the keratoconus gene. The results suggest that the causative gene in keratoconus is located within the 16q22.3-q23.1 chromosomal region.

Adolescent↗
PubMed1979

Fine structure of sensory nerves in the rat cornea: an experimental nerve degeneration study.

Fine structure of nerve fibres and terminals in the rat cornea was studied after fixation with glutaraldehyde and osmium tetroxide or with potassium permanganate. Superior cervical and/or ciliary ganglionectomy as well as ophthalmic neurotomy were performed in order to verify the origin of the corneal nerves. In the control corneas axon profiles with granular vesicles were observed in the stroma. Other axons containing agranular vesicles were found both in the stroma and in the epithelium. Superior cervical ganglionectomy abolished only the axons containing granular vesicles but did not affect the corneal blink reflex. Ciliary ganglionectomy induced more conspicuous changes in both stromal and intraepithelial axon profiles. The corneal reflex was, however, normal. Some of the axons containing agranular vesicles also showed degenerative changes. However, intact axon profiles with agranular vesicles were seen even after extirpation of both the ciliary ganglion and the superior cervical ganglion (combined autonomic ganglionectomy). On the other hand, all axon profiles with agranular vesicles disappeared after coagulation of the ophthalmic and maxillary nerves (sensory denervation). This operation destroyed most of the axons in all parts of the cornea and abolished the blink reflex. It is concluded that the axon profiles in the corneal epithelium, containing agranular vesicles, are sensory axons of the trigeminal nerve. A possible role of the vesicles in sensory nerves is discussed.

Animals↗