CHO cells expressing the human insulin receptors (IR) were used to evaluate the effect of the potent farnesyltransferase inhibitor, manumycin, on insulin antiapoptotic function. Cell treatment with manumycin blocked insulin's ability to suppress pro-apoptotic caspase-3 activity which led to time-dependent proteolytic cleavage of two nuclear target proteins. The Raf-1/MEK/ERK cascade and the serine/threonine protein kinase Akt are two survival pathways that may be activated in response to insulin. We tested the hypothesis that inhibition of farnesylated Ras was causally related to manumycin-induced apoptosis and showed that the response to manumycin was found to be independent of K-Ras function because membrane association and activation of endogenous K-Ras proteins in terms of GTP loading and ERK activation were unabated following treatment with manumycin. Moreover, blocking p21Ras/Raf-1/MEK/ERK cascade by the expression of a transdominant inhibitory mSOS1 mutant in CHO-IR cells kept cells sensitive to the antiapoptotic action of insulin. Insulin-dependent activation of Akt was blocked by 4 h treatment with manumycin (P < 0.01), a kinetic too rapid to be explained by Ras inhibition. This study suggests that the depletion of short-lived farnesylated proteins by manumycin suppresses the antiapoptotic action of insulin at least in part by disrupting Akt activation but not that of the K-Ras/Raf-1/ERK-dependent cascade.

Alkyl and Aryl Transferases↗

PubMed2000

Insulin-induced Ca(2+) entry in hepatocytes is important for PI 3-kinase activation, but not for insulin receptor and IRS-1 tyrosine phosphorylation.

Insulin produces an influx of Ca(2+) into isolated rat hepatocyte couplets that is important to couple its tyrosine kinase receptor to MAPK activity (Benzeroual et al., Am. J. Physiol. 272, (1997) G1425-G1432. In the present study, we have examined the implication of Ca(2+) in the phosphorylation state of the insulin receptor (IR) beta-subunit and of insulin receptor substrate-1 (IRS-1), as well as in the stimulation of PI 3-kinase activity in cultured hepatocytes. External Ca(2+) chelation (EGTA 4 mM) or administration of Ca(2+) channel inhibitors gadolinium 50 microM or nickel 500 microM inhibited insulin-induced PI 3-kinase activation by 85, 50 and 50%, respectively, whereas 200 microM verapamil was without effect. In contrast, the insulin-induced tyrosine phosphorylation of IR beta-subunit and of IRS-1 was not affected by any of the experimental conditions. Our data demonstrate that the stimulation of PI 3-kinase activity by the activated insulin receptor, but not the phosphorylation of IR beta-subunit and IRS-1, requires an influx of Ca(2+). Ca(2+) thus appears to play an important role as a second messenger in insulin signaling in liver cells.

Animals↗

PubMed2000

Induction of cataracts of varying degrees of hardness in human eyes obtained postmortem for cataract surgeon training.

PURPOSE: To report a method of inducing nuclear cataracts of varying degree of hardness using intralenticular injection of the Karnovsky solution. METHODS: Twelve postmortem human eyes were prepared according to the Miyake-Apple posterior video technique. After performing capsulorhexis and hydrodissection/delineation, 0.2 ml of the Karnovsky solution was injected into the lens nucleus of the eyes. Manual extracapular cataract extraction techniques and various two-handed phacoemulsification maneuvers were then practiced at various time intervals after the injection. RESULTS: Uniform nuclear cataracts with varying degrees of hardness were successfully induced in all globes. The optimum dose of the Karnovsky solution and time for induction of nuclear hardness of grade 3 (ideal for practicing two-handed phacoemulsification maneuvers) were 0.2 ml and 15 minutes, respectively. CONCLUSION: The Karnovsky solution can be successfully used for the induction of hard, uniform nuclear cataracts for learning/practicing extracapular cataract extraction and phacoemulsification.

Cataract↗

PubMed2000

Mitomycin-C and vernal conjunctivitis.

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Administration, Topical↗

PubMed2000

Postoperative deposition of calcium on the surfaces of a hydrogel intraocular lens.

OBJECTIVE: To report clinical, pathological and histochemical features of 5 Hydroview intraocular lenses (IOLs) explanted from five patients who had visual disturbances caused by postoperative deposits on the lens surfaces. DESIGN: Noncomparative small case series with clinicopathologic and histochemical correlations. PARTICIPANTS: Five hydrophilic IOLs explanted from five different patients. All patients presented with decreased visual acuity and glare circa 12 months after uneventful phacoemulsification and IOL implantation, associated with a red-brown granularity on the optical surfaces of the IOLs. METHODS: The lenses were explanted, fixed in buffered formaldehyde and examined by gross and light microscopy. MAIN OUTCOME MEASURES: Staining of the IOLs with 1% alizarin red and with the von Kossa method (both stains for calcium). Two additional IOLs were also stained and included as controls. RESULTS: The optical surfaces of all five IOLs were covered by a layer of irregular granular deposits, composed of multiple fine, translucent spherical-ovoid granules. The deposits stained positive for calcium in all cases. No deposit or positive staining was observed on the IOLs haptics. Staining of the control IOLs was also negative. CONCLUSION: This is the first histopathological report of calcified deposits on the surfaces of this hydrogel IOL model. Further studies on other similar cases with this lens should be done to determine the incidence and possible mechanisms of this phenomenon.

Aged↗

PubMed2000

Anterior capsule opacification: a histopathological study comparing different IOL styles.

OBJECTIVE: To compare the degree of anterior capsule opacification (ACO) in human eyes obtained post-mortem containing various rigid and foldable posterior chamber intraocular lens (PC-IOL) designs. DESIGN: Comparative autopsy tissue study with clinicopathologic correlations. MATERIALS: Four hundred sixty human globes containing the following PC-IOL styles were analyzed: (1) one-piece polymethylmethacrylate (PMMA) optic-PMMA haptic (n = 50), (2) one-piece silicone-plate IOL, large hole (n = 40), (3) one-piece silicone-plate IOL, small hole (n = 67), (4) three-piece PMMA optic-PMMA/Prolene haptic (n = 51), (5) three-piece acrylic optic-PMMA haptic (n = 96), (6) three-piece silicone optic-PMMA haptic (n = 24), (7) three-piece silicone optic-polyimide haptic (n = 40), and (8) three-piece silicone optic-prolene haptic (n = 92). TESTING: The globes were sectioned in the equatorial plane for gross examination and then processed through paraffin; sectioned, and stained with hematoxylin-eosin, periodic acid-Schiff, and Masson's trichrome stains; and examined by light microscopy. MAIN OUTCOME MEASURES: Anterior capsule opacification was scored in each eye by grading the histologic sections from 0 to III, according to the amount (thickness) of proliferative tissue and cells measured in sagittal sections on the inner surface of the anterior capsule at the capsulorhexis margin. RESULTS: The difference among the eight groups was significant (P < 0.0001). Mean ACO scores were highest with the large and small hole one-piece silicone-plate lenses (1.77 +/- 0.86 and 1.28 +/- 0.77, respectively). The lowest mean score was observed in the group of three-piece acrylic optic-PMMA haptics lenses (0.51 +/- 0.52). CONCLUSIONS: Our results confirm previous clinical observations that the rate of ACO is relatively high with plate-haptic silicone IOLs. The lowest rate was noted with the three-piece acrylic optic-PMMA haptic IOL. The IOL design and IOL material are significant factors in the development of ACO.

Cataract↗

PubMed2000

Dye-enhanced cataract surgery. Part 1: anterior capsule staining for capsulorhexis in advanced/white cataract.

PURPOSE: To evaluate anterior capsule staining using 3 dyes to perform continuous curvilinear capsulorhexis (CCC) in postmortem human eyes with advanced/white cataract. SETTING: Center for Research on Ocular Therapeutics and Biodevices, Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina, USA. METHODS: In experimental closed-system surgery, CCC was performed in 12 postmortem human eyes with cataract after the anterior capsule was stained with 3 capsule dyes (fluorescein sodium 2%, indocyanine green [ICG] 0.5%, and trypan blue 0.1%). Two commonly used techniques for capsule staining were also compared: staining within an air bubble and intracameral subcapsular injection of dye. RESULTS: In all globes, CCC was uneventful using the 3 dyes and with both techniques. With the intracameral subcapsular injection, the dye remained trapped in the subcapsular space in contact with the posterior surface of the anterior capsule, allowing enough time to perform any maneuver. The staining provided by ICG, at the concentration used, was slightly superior to that of the other dyes. Leakage of fluorescein sodium into the vitreous cavity was seen using the Miyake-Apple posterior video/photographic technique. CONCLUSION: Intracameral subcapsular injection of ICG allowed the easiest recognition of the capsular flap by staining the posterior surface of the anterior capsule and without leaking into the vitreous cavity.

Cadaver↗

PubMed2000

Dye-enhanced cataract surgery. Part 3: posterior capsule staining to learn posterior continuous curvilinear capsulorhexis.

PURPOSE: To evaluate the use of 2 dyes for staining the posterior capsule to enhance visualization during posterior continuous curvilinear capsulorhexis (PCCC). SETTING: Center for Research on Ocular Therapeutics and Biodevices, Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina, USA. METHODS: Indocyanine green 0.5% (ICG) and trypan blue 0.1% were used to stain the posterior capsule for performing PCCC in 8 human eyes obtained postmortem. The eyes were prepared according to the posterior video technique of Miyake-Apple. After CCC and hydrodissection/delineation were performed, the nucleus was carefully delivered by hydroexpression. Cortical cleanup was completed by irrigation/aspiration. The posterior capsule was stained by instilling 1 microdrop of ICG (4 eyes) or trypan blue (4 eyes) in the capsular bag. A PCCC was then performed by 2 independent surgeons (4 eyes/surgeon) not familiar with this technique. Each surgeon also performed PCCC in 2 eyes without the use of dye. RESULTS: Both dyes facilitated the performance of PCCC after staining the otherwise transparent posterior capsule. It was easier to differentiate the stained posterior capsule flap from the underlying transparent anterior hyaloid phase of the vitreous. In addition, optic capture of an intraocular lens, with or without anterior vitrectomy, was easily accomplished because of the enhanced visualization by posterior capsule staining. CONCLUSIONS: Posterior capsule staining can be successfully used to learn and perform the PCCC procedure, combined with optic capture, anterior vitrectomy, or both.

Cadaver↗

PubMed2000

Dye-enhanced cataract surgery. Part 2: learning critical steps of phacoemulsification.

PURPOSE: To use capsule dyes to enhance visualization to learn and perform various critical steps of the phacoemulsification procedure in a laboratory setting using postmortem human eyes. SETTING: Center for Research on Ocular Therapeutics and Biodevices, Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina, USA. METHODS: Indocyanine green 0.5% (ICG) and trypan blue 0.1% were used to enhance visualization for performing critical steps of phacoemulsification in 8 human eyes obtained postmortem. All eyes were prepared according to the Miyake-Apple posterior video technique. After a continuous curvilinear capsulorhexis (CCC) was made, the dyes were used to enhance visualization for hydrodissection, hydrodelineation, and various maneuvers for nuclear emulsification and cortical cleanup. In 8 eyes, all the aforementioned steps were performed without using dye (control group). RESULTS: Both dyes helped enhance visualization during the critical steps of phacoemulsification. The use of a dye solution instead of balanced salt solution (BSS) for hydrodissection/delineation helped localize the complete plane of cleavage between the capsule and cortex and the nucleus-epinucleus complex. During various nuclear sculpting maneuvers, the use of dye helped visualization of the position of the phaco tip and its relation to the posterior capsule. It also helped localize remaining cortical fibers, facilitating complete cleaning of the capsular bag. CONCLUSIONS: Dye-enhanced cataract surgery was useful in learning various critical steps of phacoemulsification in a wet laboratory setting.

Cadaver↗

PubMed2000

Opacification of piggyback IOLs associated with an amorphous material attached to interlenticular surfaces.

PURPOSE: To report the pathological and ultrastructural features and interval surgical management of an atypical case of opacification between piggyback intraocular lenses (IOLs). SETTING: Center for Research on Ocular Therapeutics and Biodevices, Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina, and Nature Coast EyeCare Institute and Surgery Center, Perry, Florida, USA. METHODS: Opacification between 2 acrylic piggyback lenses was observed 16 months after implantation, with decreased best corrected visual acuity and a hyperopic shift. Elschnig pearls were observed in the peripheral interface between the lenses, and the central interface was occupied by an amorphous material. The pearls were surgically aspirated, but attempts to remove the central material were unsuccessful. The lenses were explanted and sent to the laboratory. Staining with hematoxylin and eosin (H&E), examination under a light microscope, and scanning electron microscopic analysis were performed. RESULTS: The surfaces of the anterior IOL were relatively clear. The amorphous material, mostly attached to the center of the anterior surface of the posterior IOL, was homogeneously stained with H&E. No cell nucleus was observed in this region. Scanning electron microscopy showed that the IOL edge presented a smooth, regular surface relatively free of deposits. The most central region was covered by an irregular layer of an amorphous compact material with some cracks, fissures, or both on its surface. CONCLUSIONS: Although the exact composition of the material between the lenses could not be established, hypotheses were advanced to understand the pathological mechanism associated with this condition. This case is different from those in previous reports of opacification composed of cortex and cells between piggyback IOLs.

Acrylates↗

PubMed2000

Deposits on the optical surfaces of Hydroview intraocular lenses.

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Biocompatible Materials↗

PubMed2000

Adhesion of fibronectin, vitronectin, laminin, and collagen type IV to intraocular lens materials in pseudophakic human autopsy eyes. Part 2: explanted intraocular lenses.

PURPOSE: To evaluate fibronectin, vitronectin, laminin, and collagen type IV adhesion to poly(methyl methacrylate) (PMMA), silicone, hydrophobic soft acrylate, and hydrogel intraocular lenses (IOLs) in human pseudophakic autopsy eyes. SETTING: Center for Research on Ocular Therapeutics and Biodevices, Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina, USA. METHODS: Thirty-two autopsy eyes containing PMMA, silicone, soft acrylate, or hydrogel IOLs were assessed. The IOLs were explanted from the capsular bag, and both sides of the IOLs were immunohistochemically stained for fibronectin, vitronectin, laminin, or collagen type IV. The number of cells on the IOL surfaces was counted. The capsular bag from 1 eye containing a soft acrylate IOL was examined for fibronectin and vitronectin. RESULTS: Hydrophobic soft acrylate IOLs had significantly more fibronectin adhering to their surfaces than PMMA (P <.01) or silicone (P <.01) IOLs, as well as more vitronectin. Silicone IOLs had more collagen type IV adhesion than the other IOLs (P <.05-.06). Collective protein adhesion differed significantly between soft acrylate IOLs and PMMA and silicone IOLs, but not between PMMA and silicone IOLs. CONCLUSIONS: The greater amount of protein on the hydrophobic soft acrylate (AcrySof(R)) IOLs seems to support an adhesive mechanism for their attachment to the capsular bag. Fibronectin and vitronectin have functional domains to bind them to lens epithelial cells and the collagenous capsule. This kind of attachment could be a true bioactive bond and may be 1 reason the PCO and neodymium:YAG capsulotomy rates are lower in eyes with a soft acrylate IOL.

Acrylic Resins↗

PubMed2000

Adhesion of fibronectin, vitronectin, laminin, and collagen type IV to intraocular lens materials in pseudophakic human autopsy eyes. Part 1: histological sections.

PURPOSE: To evaluate fibronectin, vitronectin, laminin, and collagen type IV adhesion to poly(methyl methacrylate) (PMMA), silicone, hydrophobic soft acrylate, and hydrogel intraocular lenses (IOLs) in pseudophakic human autopsy eyes. SETTING: Center for Research on Ocular Therapeutics and Biodevices, Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina, USA. METHODS: Thirty-eight autopsy eyes containing PMMA, silicone, hydrophobic acrylate, or hydrogel IOLs were assessed. Histological sections were prepared from each eye, and immunohistochemical analyses were performed for fibronectin, vitronectin, laminin, and collagen type IV. One hundred fifty-two specimens were analyzed. RESULTS: A sandwich-like structure (anterior or posterior capsule/fibronectin/1 cell layer/fibronectin/IOL surface) was seen in 12 of 14 autopsy eyes with soft acrylate IOLs, 3 of 10 with a PMMA IOL (P =.0094), 1 of 10 with a silicone IOL (P =.0022), and 0 of 4 with a hydrogel IOL (P =. 0041). The thicker fibrocellular tissue on the inner surface of the anterior or posterior capsule that was in contact with silicone IOLs was lined with collagen type IV. Vitronectin and laminin were not found at the fibrocellular tissue-IOL interface in any specimen. CONCLUSIONS: This study seems to confirm the sandwich theory of posterior capsule opacification in eyes with an IOL and suggests that fibronectin may be the major extracellular protein responsible for the attachment of hydrophobic soft acrylate (AcrySof(R)) IOLs to the capsular bag. This may represent a true bioactive bond between the IOL and lens epithelial cells or between the IOL and the capsular bag and may be one reason the PCO and neodymium:YAG capsulotomy rates are lower in eyes with a soft acrylate IOL.

Biocompatible Materials↗