Juergen Kampmeier

Effect of growth factors on proliferation and expression of growth factor receptors in a human lens epithelial cell line.

PURPOSE: To investigate the effect of basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), insulin‐like growth factor 1 (IGF‐1), and transforming growth factor beta 2 (TGFβ2) on proliferation of a human lens epithelial cell line (HLEC‐SRA 01/04); the effect of bFGF and TGFβ2 on proliferation of human lens epithelial cells (HLECs); and the expression of bFGF, EGF, IGF‐1, and TGFβ2 receptors in an HLEC‐SRA 01/04 cell line. SETTING: Department of Ophthalmology, University of Ulm, Ulm, Germany. METHODS: Both HLEC and HLEC‐SRA 01/04 were treated with 1 to 50 ng/mL bFGF and TGFβ2. Additionally, HLEC‐SRA 01/04 were cultured with EGF and IGF‐1 at a concentration of 1 to 50 ng/mL for 48 hours in the presence of [3H]‐thymidine. In all experiments, untreated serum‐free negative controls were used. (3H)‐thymidine incorporation as a direct measure of lens epithelial cell proliferation was assessed by liquid scintillation counting. The expression of bFGF, EGF, IGF‐1, and TGFβ2 receptors in HLEC‐SRA 01/04 were analyzed by reverse transcriptase polymerase chain reaction (RT‐PCR). Statistical analysis was performed using the 2‐sample t test for the means. RESULTS: Proliferation of HLECs was dose dependently induced by bFGF and TGFβ2, showing maximum effects at 10 ng/mL (P= .0003) and at 50 ng/mL (P<.0001), respectively. Proliferation of HLEC‐SRA 01/04 was also induced by bFGF, showing slight but significant effects (P<.03). Additionally, HLEC‐SRA 01/04 proliferation was dose‐dependently induced by EGF with a maximum effect at 5 ng/mL (P<.01), IGF‐1 with a maximum effect at 5 ng/mL (P<.0001), and TGFβ2 with a maximum effect at 10 ng/mL (P<.0001) compared with the control. The RT‐PCR analysis revealed bFGF, EGF, IGF‐1, and TGFβ2 receptor expression in the HLEC‐SRA 01/04 cell line. CONCLUSIONS: The data showed that bFGF and TGFβ2 are strong mitogens for HLEC. The HLEC‐SRA 01/04 cell line derived from HLEC reacted to growth factors, with cell proliferation only to a lesser extent. Such quiescence of these cells, when compared with cells in primary culture, cannot be explained by the lack of respective receptors for growth factors. Further investigation of growth factor‐induced responses of both cell types will provide new insight into the proliferative processes involved in postoperative secondary cataract formation.

Effect of extracellular matrix on proliferation and differentiation of porcine lens epithelial cells

BackgroundProliferation and differentiation of lens epithelial cells (LECs) are important mechanisms of secondary cataract formation. After extracapsular cataract extraction the extracellular matrix (ECM) around the remaining LECs is altered compared with the intact lens. This study investigated the effects of different ECMs on cell proliferation and alpha-smooth muscle actin (α-SMA) expression, a marker for myofibroblasts, in cultured porcine LECs.MethodsPorcine LECs were cultured for 3 days (cell proliferation assay) or 4 days (α-SMA expression) on wells and glass cover slips, respectively, coated with laminin, fibronectin, type I collagen or type IV collagen. LECs cultured on uncoated wells or cover slips served as control. Proliferative response was measured by [3H]-thymidine incorporation into DNA. α-SMA was detected immunocytochemically with a mouse monoclonal antibody, and the relative numbers of α-SMA-positive cells were calculated. Statistical analysis was performed using Student’s unpaired t-test.ResultsCell proliferation was significantly increased by coating with fibronectin (10,320.5±6,073 counts per minute; p<0.0001) (mean ± SD), type I collagen (12,507.3±3,914.2 CPM; p<0.0001) and type IV collagen (9,591.4±4,088 CPM; p<0.0001) compared with control (1,876.5±998 CPM), whereas coating with laminin had no effect (1,760.8±812.6 CPM; p=0.7271). The ratio of α-SMA-positive LECs cultured on uncoated cover slips for a period of 4 days was 12.2±3.51%. This ratio was significantly increased by coating with fibronectin (24.3±4.56%; p=0.0001) and type I collagen (21.2±8.48%; p=0.0142). Coating with laminin (9.8±3.67%; p=0.1682) and type IV collagen (9.0±7.09 %; p=0.2491) slightly decreased α-SMA expression, but this effect was not statistically significant.ConclusionsFibronectin and type I collagen stimulated both cell proliferation and α-SMA expression in cultured porcine LECs. Because fibronectin and type I collagen are not normally present in the adult lens, their possible introduction into the lens capsule after cataract surgery may play a critical role in the development of posterior capsule opacification.

Comparison of free-running vs. Q-switched Er:YAG laser photorefractive keratectomy (scanning mode) in swine eyes.

PURPOSEnTo examine morphology in plane and incisional corneal ablation (in vitro) induced by an Er:YAG laser (2.94 microm) in two modes: free-running and q-switched.nnnMETHODSnSequences of different fluences in each mode were applied to freshly enucleated pig eyes. Parameters of free-running mode were: pulse length 50 micros, fluences 1.21 to 4.77 J/cm2, frequency 80 Hz, spot size 500 microm FWHM, hexagonal spot shape. Parameters of q-switched mode were: pulse length 200 ns, fluences 0.79 to 2.33 J/cm2, frequency 20 Hz, spot size 500 microm FWHM, round spot shape.nnnRESULTSnHistology showed thermal damage of 10 to 25 microm in depth caused by the free-running mode compared with 4.5 to 7.5 microm by the q-switched mode. In both gross photography and scanning electron microscopic examination, the surface was more homogeneous and smoother in the q-switched mode.nnnCONCLUSIONSnDepending on the different application modes, both laser systems could be used for a defined corneal ablation in photorefractive keratectomy. At present, results using the Er:YAG laser are not as favorable as with the excimer laser.

An intraocular micro light-emitting diode device for endo-illumination during pars plana vitrectomy

Purpose: Development of a new, fiber-free, single-use endo-illuminator for pars plana vitrectomy as a replacement for fiber-based systems with external light sources. The hand-guided intraocularly placed white micro light-emitting diode is evaluated for its illumination properties and potential photochemical and thermal hazards. Methods: A micro light-emitting diode was used to develop a single-use intraocular illumination system. The light-source-on-tip device was implemented in a prototype with 23G trocar compatible outer diameter of 0.6 mm. The experimental testing was performed on porcine eyes. All calculations of possible photochemical and thermal hazards during the application of the intraocular micro light-emitting diode were calculated according to DIN EN ISO 15007–2: 2014. Results: The endo-illuminator generated a homogeneous and bright illumination of the intraocular space. The color impression was physiologic and natural. Contrary to initial apprehension, the possible risk caused by inserting a light-emitting diode into the intraocular vitreous was much smaller when compared to conventional fiber-based illumination systems. The photochemical and thermal hazards allowed a continuous exposure time to the retina of at least 4.7 h. Conclusion: This first intraocular light source showed that a light-emitting diode can be introduced into the eye. The system can be built as single-use illumination system. This light-source-on-tip light-emitting diode–endo-illumination combines a chandelier wide-angle illumination with an adjustable endo-illuminator.

Photorefraktive Keratektomie@@@Free-running versus q-switched Er:YAG-laser (scanning mode) in photorefractive keratectomy: Freilaufender vs. gütegeschalteter Er:YAG-Laser (Scanning Modus)

ZusammenfassungFragestellung: Vergleichende morphologische Untersuchung zur planen photorefraktiven Keratektomie mit dem gütegeschalteten (200 ns) und dem freilaufenden (50 μs) Er:YAG-Laser (2,94 μm).nMaterial und Methode: An frisch enukleierten Schweineaugen wurden im Scanning-Modus Pulse verschiedener Energiedichten appliziert. Freilaufende Parameter: Energiedichte: 1,21–4,77 J/cm2, Frequenz: 80 Hz, Spotgröße: 500 μm Halbwertsbreite, hexagonale Spotgeometrie. Gütegeschaltete Parameter: Energiedichte: 0,79–2,33 J/cm2, Frequenz: 20 Hz, Spotgröße: 500 μm Halbwertsbreite, runde Spotgeometrie.nErgebnisse: Histologisch zeigt sich eine thermische Nekrosezone von 10–25 μm im freilaufenden Modus im Vergleich zu 4,5–7,5 μm im gütegeschalteten Modus. In der Rasterelektronenmikroskopie findet sich im gütegeschalteten Modus eine vergleichsweise homogenere und glattere korneale Oberfläche.nSchlußfolgerung: Abhängig von verschiedenen Applikationsformen können beide Lasersysteme für einen definierten kornealen Abtrag in der photorefraktiven Chirurgie eingesetzt werden. Jedoch kann zum jetzigen Zeitpunkt die Oberflächenqualität von Excimer-Laser-Systemen nicht erreicht werden.SummaryBackground: Examination of morphology in plane corneal ablation (in vitro) induced by an Er:YAG-laser (2.94 μm) in two modes: free-running (50 μs) and q-switched (200 ns).nMethods: Sequences of different fluences in each mode were applied to freshly enucleated swine eyes. Parameters of free-running mode: fluences 1.21–4.77 J/cm2, frequency 80 Hz, spot size 500 μm FWHM, hexagonal spot shape. Parameters of q-switched mode: fluences 0.79–2.33 J/cm2, frequency 20 Hz, spot size 500 μm FWHM, round spot shape.nResults: Histology showed thermal damage of 10–25 μm in depth caused by the free-running mode compared with 4.5–7.5 μm by the q-switched mode. In both gross photography and scanning electron microscopic examination, the surface was found to be more homogeneous and smoother in the q-switched mode.nConclusions: Depending on the different application modes, both laser systems could be used for a defined corneal ablation in photorefractive keratectomy. However, at the moment, results using the Er:YAG laser are not as favorable as with the excimer laser.