Sai-ichi Tanaka

Fibrotic disorders in the eye: targets of gene therapy.

Fibrotic diseases, e.g., cutaneous and corneal scarring, keloids, and liver and lung fibrosis, etc., are characterized by appearance of myofibroblasts, the key player of the fibrogenic reaction, and excess accumulation of extracellular matrix with resultant tissue contraction and impaired functions. Inflammatory/fibrogenic growth factors/cytokines produced by injured tissues play a pivotal role in fibrotic tissue formation. Ocular tissues are also susceptible to fibrotic diseases. In this article, the pathogenesis of such fibrotic disorders in the eye, i.e., scarring in the cornea and conjunctiva, post-cataract surgery fibrosis of the lens capsule and proliferative vitreoretinopathy are reviewed. Focus is put on the roles of myofibroblast and signals activated by the fibrogenic cytokine, transforming growth factor beta. Modulation of signal transduction molecules, e.g., Smad and mitogen-activated protein kinases, by gene transfer and other technology is beneficial and can be an important treatment regiment to overcome (prevent or treat) these diseases.

Lens epithelial cell outgrowth and matrix formation on intraocular lenses in rabbit eyes

Purpose: To evaluate the presence and distribution of lens epithelial cells (LECs) and extracellular matrix on intraocular lenses (IOLs) implanted in the capsular bag in rabbit eyes. Setting: Department of Ophthalmology, Wakayama Medical College, Wakayama, Japan. Methods: Five adult albino rabbits had phacoemulsification and IOL implantation in both eyes. Two or 11 months later, the animals were killed by intravenous pentobarbital. The IOLs were removed and observed under scanning and transmission electron microscopy. Results: In addition to the macrophages and giant cells on the IOLs, all eyes had a monolayer of flattened cells growing out from the residual anterior lens capsule and a fibrous meshwork of extracellular matrix. Unlike those of a macrophagic origin, these cells had no central cytoplasmic elevation of nuclei and few cell surface microvilli and were considered to be proliferating LECs. Conclusion: Lens epithelial cells are involved in the eye’s cellular reaction to IOLs and in the formation of extracellular matrix on IOLs. Further study of LEC behavior on IOLs should be done to improve IOL biocompatibility.

Epithelial-mesenchymal transition as a therapeutic target for prevention of ocular tissue fibrosis.

Fibrotic diseases are characterized by the appearance of myofibroblasts, the key cell type involved in the fibrogenic reaction, and by excess accumulation of extracellular matrix with resultant tissue contraction and impaired function. Myofiborblasts are generated by fibroblast-myofibrobalst conversion, and in certain tissues through epithelial-mesenchymal transition (EMT), a process through which an epithelial cell changes its phenotype to become more like a mesenchymal cell. Although inflammatory/fibrogenic growth factors/cytokines produced by injured tissues orchestrate the process of EMT, transforming growth factor beta (TGFbeta) is believed to play a central role in the process. Unlike fibrotic lesions in kidney or other tissues where myofibroblasts are generated from both fibroblasts and epithelial cells, fibrotic lesions in the eye crystalline lens are derived only from lens epithelial cells without contamination of fibroblast-derived myofibroblasts. Thus, this tissue is suitable to investigate detailed mechanisms of EMT and subsequent tissue fibrosis. EMT in retinal pigment epithelium is involved in the development of another ocular fibrotic disease, proliferative vitreoretinopathy, a fibrosis in the retina. EMT-related signal transduction cascades, i. e., TGFbeta/Smad, are a target to prevent or treat unfavorable ocular tissue fibrosis, e. g., fibrotic diseases in the crystalline lens or retina, as well as possibly in other organs.

Extracellular matrix on intraocular lenses

We examined the distribution of extracellular matrix components in the proteinaceous matrix on explanted IOLs using immunohistochemistry to clarify the nature of proteinaceous deposits on the surfaces of IOLs. We examined 15 polymethylmethacrylate (PMMA) IOLs and one silicone IOL explanted from patients. The IOLs were immunostained for cellular fibronectin, types I and IV collagen, and vitronectin. Various amounts of fibronectin, types I and IV collagen, and vitronectin were detected in the extracellular matrix on all IOLs with cellular deposits. Types I and IV collagen and cellular fibronectin were present in the extracellular matrix and were probably the products of cells adhering to the IOLs. Vitronectin in the fibrous extracellular matrix appeared to represent an adsorption material derived from the aqueous humor. These proteinaceous deposits associated with cellular deposits indicate unsuccessful formation of granulation tissue and could influence IOL biocompatibility.

Deposition of extracellular matrix on intraocular lenses in rabbits : an immunohistochemical and transmission electron microscopic study

Abstract• Background: We examined by transmission electron microscopy the accumulation of extracellular matrix on intraocular lenses (IOLs) implanted experimentally into rabbit eyes, and evaluated the immunolocalization of such extracellular matrix components as collagen types I, III, and IV, and cellular fibronectin on these IOLs. • Methods: Phacoemulsification and aspiration of the crystalline lens were performed and an IOL was implanted into the capsular bag of each eye of each of 16 adult albino rabbits under general anesthesia. After up to 12 weeks, the animals were killed and the IOLs were removed. Specimens were processed for transmission electron microscopy or for immunohistochemical detection collagen types I, III, and IV, and cellular fibronectin. • Results: Transmission electron microscopy revealed an accumulation of extracellular matrix between the residual anterior lens capsule and the surface of an IOL explanted 4 weeks after surgery. Collagen types I and III and cellular fibronectin were detected immunohistochemically on each IOL in association with cellular deposits. Type IV collagen-immunoreactive matrix was not seen on the optic portion, but was detected on the haptic portion of one of six IOLs examined. • Conclusion: Each component of the extracellular matrix that is deposited on the IOL supplies scaffolding for the adhesion and proliferation of cells. These components are considered to be produced by cells such as lens epithelial cells and macrophages that adhere to the IOL surface.

Matrix metalloproteinases and tissue inhibitors of metalloproteinases of fibrous humans lens capsules with intraocular lenses

Purpose. We located immunohistochemically the matrix metalloproteinases (MMP) -1, -2, -3 and -9 and the tissue inhibitors of matrix metalloproteinases (TIMP) -1 and -2 in the fibrous capsule of patients with intraocular lenses (IOLs). Methods. During vitreoretinal surgery in 10 patients we obtained post-cataract surgery lens capsules with or without an IOL. The mean interval between the previous cataract operation and the extraction of the specimens was 35.2 months (range: 2-120 months). Circular sections of the anterior capsule with lens epithelial cells (LECs) were also obtained during cataract surgery. Specimens were processed for immunohistochemical identification of MMPs and TIMPs by light microscopy. Results. While all the members of MMPs and TIMPs were not detected in the normal anterior capsules, they were detected in the ECM and/or LECs on the lens capsules extracted within 18 months after IOL implantations in all of the 4 patients, but were not observed in specimens obtained 18 months or longer postoperatively. In LECs of 1 capsule specimen 10 years postoperatively, MMP-1, but not other MMPs and TIMPs, was detected. Conclusions. MMPs and TIMPs were detected in the ECM and/or LECs on post-cataract surgery capsules. These proteins may be remodeling the newly deposited ECM and regulating LEC behavior on residual lens capsules in the early phase of healing after cataract surgery.

Immunolocalization of hyaluronan and CD44 in quiescent and proliferating human lens epithelial cells.

Purpose: To investigate the role of hyaluronan and its receptor CD44 in capsular repair, their localization in opacified human posterior capsules and in lens epithelial cells (LECs) was assayed. Setting: Research laboratory, Department of Ophthalmology, Wakayama Medical College, Japan. Methods: In 8 patients, circular sections of the anterior capsules obtained during cataract surgery, the extracted crystalline lens, and the opacified lens capsules resulting from intraocular lens implantation were examined immunohistochemicaliy f to detect hyaluronan or CD44. Results: Both hyaluronan and CD44 were found in the extracellular matrix that accumulated on the inner surface of the capsular bags. In contrast, LECs exhibited immunoreactivity to CD44 but not to hyaluronan. Conclusions: The cell surface antigen CD44 was expressed ubiquitously by‐ LECs. Expression of its ligand hyaluronan by proliferating LECs suggests that this glycosaminoglycan may be important in the development of posterior capsule opacification.

Light and scanning electron microscopy of rabbit lens capsules with intraocular lenses

Purpose: To examine postoperative changes in the lens capsules of rabbit eyes after phacoemulsification and aspiration of the crystalline lens and implantation of posterior chamber intraocular lenses (IOLs) using light and scanning electron microscopy. Setting: Research Laboratory, Department of Ophthalmology, Wakayama Medical College, Japan. Methods: The crystalline lens was emulsified and aspirated and an IOL implanted in the capsular bag or ciliary sulcus of each eye in adult albino rabbits under general anesthesia. Animals were killed after 4 weeks, and the lens capsules were removed. The specimens were observed under phase‐contrast microscopy and processed for light and scanning electron microscopy. Results: Phase‐contrast microscopy revealed presumed lens epithelial cells (LECs) on the central posterior capsules in association with regenerating lenticular fibers and Elschnig pearls in the peripheral capsules. Scanning electron microscopy showed the accumulation of fibrous extracellular matrix on the surface of the posterior capsule in eyes in which the IOL was implanted in the ciliary sulcus. Deposition of packed material attached to the surface of IOLs and of Soemmering’s ring were observed in eyes with in‐the‐bag IOL fixation. At a higher magnification, a parallel arrangement of lenticular fibers was seen in the regenerated lens structure on posterior capsules. An identical structure was observed under light microscopy. Outgrowth of presumed LECs from residual anterior lens capsules and adhesion of macrophages and giant cells were observed on the IOL surface. Conclusion: Two types of postoperative changes were observed in lens capsules after implantation of IOLs: accumulation of fibrous extracellular matrix and newly formed lenticular fibers. These changes are attributed to the proliferation of LECs and can induce posterior capsule opacification after IOL implantation.

Subretinal administration of tissue-type plasminogen activator to speed the drainage of subretinal hemorrhage

Abstract • Background: Bleeding into the subretinal space in the vicinity of the macula is associated with age-related macular degeneration or retinal arterial macroaneurysm. The prognosis for restoration of vision is poor in the presence of blood clots. • Methods: Using a simple device composed of three disposable syringes we injected tissue-type plasminogen activator (tPA) into the subretinal space during conventional vitrectomy in six patients to assist the draining of subretinal clots. • Results: Four of six patients recovered their visual acuity postoperatively, while visual acuity in the other patients was stabilized. • Conclusion: Early drainage of subretinal hemorrhage assisted by the introduction of tPA into the subretinal space led to uncomplicated surgery and favorable postoperative results.

Transforming Growth Factor β Signal Transduction: A Potential Target for Maintenance/Restoration of Transparency of the Cornea

Maintenance of the transparency and regular shape of the cornea are essential to the normal vision, whereas opacification of the tissue impairs vision. Fibrogenic reaction leading to scarring in an injured cornea is characterized by appearance of myofibroblasts, the key player of the fibrogenic reaction, and excess accumulation of fibrous extracellular matrix. Inflammatory/fibrogenic growth factors/cytokines produced by inflammatory cells play a pivotal role in fibrogenic response. Signaling systems involved in myofibroblast formation and fibrogenesis are activated by various growth factors, i.e., transforming growth factor &bgr; or others. Modulation of transforming growth factor &bgr; signal transduction molecules, e.g., Smad and mitogen-activated protein kinases, by gene transfer and other technology provides a new concept of prevention/treatment of unfavorable fibrogenesis in the cornea.