Biomedical subjects

Zhichong Wang

Publications and source records attributed to Zhichong Wang.

9 recordsLinked to original sources

PubMed2007

Establishment of a corneal epithelial cell line spontaneously derived from human limbal cells.

The objective of this study was to establish a spontaneously derived human corneal epithelial cell line from a normal human limbus that retains differentiation potential and proliferative properties under continuous cell culture. After 50 passages of epithelial cells obtained from human limbal tissue a cell line spontaneously emerged. The immortalized cells showed a cobblestone appearance and displayed dense microvilli on their apical cell surface membrane. Colony forming efficiency was 5-6% and population doubling time was 19.6 h. In the mRNA level, cytokeratin (CK) 3 and 12 were detected in this cell line. In the protein level, the cells expressed CK3, CK12, CK14, CK19, vimentin, and some other proteins such as F-actin and beta-tubulin and beta(1)-integrin. They lacked p63. The immortalized cells had a heteroploid karyotype, but did not exhibit tumorigenic features. When cultured on an air-liquid interface the cells could form stratified multilayer epithelia. In summary, all these results indicated that a new human corneal epithelial cell line was spontaneously established from normal limbal tissue through serial culture. This cell line would be useful for studies of corneal epithelial biology and reconstructive corneal tissue engineering.

Blotting, Western↗

PubMed2006

Expression of protein kinase C isoforms in cultured human retinal pigment epithelial cells.

BACKGROUND: Protein kinase C (PKC) is involved in both physiological and pathophysiological processes and plays an important role in signal transduction. The present studies were designed to examine the 12 isoforms (PKCalpha, PKCbetaI, PKCbetaII, PKCgamma, PKCdelta, PKCepsilon, PKCeta, PKCtheta, PKCmu, PKCxi, PKClambda and PKCiota) of PKC expressed in cultured human retinal pigment epithelium (RPE) cells. METHODS: Human RPE cells were investigated for 12 PKC isoforms at the mRNA, protein and cellular levels by reverse transcription (RT)-PCR, Western blot analysis and laser scanning confocal microscope (LSCM), respectively. RESULTS: RT-PCR and Western blot analyses showed similar results for specific PKC isoforms in that both revealed that PKCalpha, PKCbetaI, PKCbetaII, PKCdelta, PKCepsilon, PKCtheta, PKCmu, PKCxi, PKClambda and PKCiota, but not PKCgamma and PKCeta, were constantly expressed in RPE cells, with the exception of PKCbetaI at the protein level. Confocal microscopy showed that ten PKC isoforms - PKCalpha, PKCbetaI, PKCbetaII, PKCdelta, PKCepsilon, PKCtheta, PKCxi, PKCiota, PKClambda and PKCmu - appeared almost exclusively in the cytoplasm of the cells. However, PKCgamma and PKCeta were not detected by staining. CONCLUSIONS: This study characterized the expression pattern of all 12 PKC isoforms and showed that ten of these (PKCalpha, PKCbetaI, PKCbetaII, PKCdelta, PKCepsilon, PKCtheta, PKCmu, PKCxi, PKClambda and PKCiota) are present in cultured human RPE cells. This identification provides the first step towards elucidating their roles in RPE cell proliferation.

Blotting, Western↗

PubMed2006

New technique for lacrimal system intubation.

PURPOSE: To propose a new technique for silicone intubation in disorders of the lacrimal system in children and adults. DESIGN: Retrospective, noncomparative case series. METHODS: The surgery was performed in 513 eyes of 426 patients from March 2003 to July 2005 in our hospital. With the help of a so-called soft probe and intranasal endoscope, silicone tubes were inserted through both puncta into nasolacrimal duct and left in place for three to six months. RESULTS: The surgery was performed successfully in all cases. No intraoperative complications were found in the procedure. Tubes were removed between 2.1 and 5.3 months (mean 3.4 months) after surgery. Mean follow-up time after tube removal was 15.1 months (range 3.3 to 27.5 months). Tube displacement occurred in 11 cases (2.14%); no other complications associated with silicone tubes, such as granuloma formation or corneal erosion, occurred. All the tubes were removed successfully. No ophthalmic injuries occurred during tube removal. CONCLUSIONS: The technique proved to be effective in both children and adults. This technique, which is atraumatic, easy to perform, time-saving, and better tolerated, may be considered as an alternative method for lacrimal system intubation.

Adolescent↗

PubMed2006

In vivo and in vitro inhibitory effect of amniotic extraction on neovascularization.

PURPOSE: To prepare amniotic extraction (AE) and to test its antiangiogenic effect in vivo and in vitro. METHODS: AE was prepared and diluted to 50, 100, and 200 microg/mL concentrations. Alkali burn-induced corneal neovascularization (NV) was produced and topically treated with different concentrations of AE or 0.1% dexamethasone for 7 days. Normal saline was used as a control. Corneal NV was visualized by heart perfusion of Chinese ink and quantified as the percentage of corneal NV area to the whole corneal area. Human umbilical vein endothelial cells (HUVECs) were primarily cultured. The effects of AE on proliferation and tube formation of HUVECs were tested by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method and in vitro angiogenesis assay. Pigment epithelium-derived factor (PEDF) in AE was detected by Western blot. RESULTS: Relative corneal NV area in the control group was 56.6% +/- 9.9%, which was significantly reduced by 50 microg/mL AE (47.6% +/- 6.9%; P = 0.043) and 200 microg/mL AE (34.3% +/- 7.8%; P < 0.001) and by 0.1% dexamethasone (21.1% +/- 1.8%; P < 0.001). HUVEC cell proliferation was significantly decreased after treatment with AE at concentrations of 50 and 100 microg/mL compared with control (P = 0.036 and 0.001, respectively). The tube formation was significantly suppressed by 100 microg/mL AE (70.03% +/- 4.35%) compared with control (100% +/- 4.84%; P = 0.002). No expression of PEDF was detected in AE. CONCLUSION: AE inhibits NV induced by alkali burn. This effect may be elicited at least in part through the inhibiting activity of blood vessel endothelial cells and is not associated with PEDF.

Amnion↗

PubMed2005

Differentiation of embryonic stem cells into corneal epithelium.

Our project was to determine whether embryonic stem (ES) cells could be induced to differentiate into corneal epithelia by superficial corneoscleral limbal stroma. To achieve this goal, ES-GFP cell line D3 was pre-induced by retinoic acid (RA). The pre-induced cells were seeded on deepithelialized superficial corneoscleral slices (SCSS) to form a monolayer, and divided into three groups. Group 1 was cultured and passaged in vitro for direct detection. Group 2 was exposed to air-liquid interfaces for 10 days and implanted into the subcutaneous layer of nude mice for 2 weeks for further induction in vivo. Group 3 was cultured in vitro without any inducing factors for control. There were no teratomas found in nude mice which were implanted with differentiated ES cells after two weeks. The differentiated cells showed an appearance of epithelia both in vitro and in vivo. Expression of CK3, P63 and PCNA was detected by immunohistochemical staining in the differentiated cells in group 1 and 2. Microvillis and zonula occludens were observed on the surface of the differentiated cells under an electron microscope. In the control group, ES cells differentiated freely without any inducing factors. Most cells were shed and formed a neuronal dendrite-like structure, and a minority of cells appeared polymorphic. These results demonstrate that ES cells can differentiate into corneal epithelia on the surface of SCSS under the controlled condition. Differentiated ES cells could be used as epithelial seeding cells for the reconstruction of ocular surface and corneal tissue engineering in the future.

Animals↗

PubMed2004

[Expression changes of protein kinase Calpha during differentiation of mouse embryonic stem cells into neuron-like cells].

PURPOSE: To study the expression of protein kinase Calpha (PKCalpha) during differentiation of mouse embryonic stem cells (ESC)into neuron-like cells in an attempt to elucidate their role in signaling. METHOD: ES-D3 cells were subjected to an 18-day induction procedure which consisted of 4 days of culture as embryoid bodies (EBs) without all-trans retinoic acid (RA) followed by 14 days of culture in the presence of RA, then the EBs were plated separately onto gelatin-coated 6-well culture dishes coated glass coverslip for immunohistochemistry, onto 100-mm-diameter bacteriological (nontissue culture) dish for Western blot assay and RT-PCR, and were cultivated and collected cells for 1, 3, 5, 7 and 14 days in the presence of RA. RESULTS: Staining of PKCalpha was strong and distributed throughout the cells, especially in the cytoplasm and membrane. Western blot analysis exhibited a band of approximately 84 kD with the antibody specific to PKCalpha. After induction of RA, PKCalpha was present in significantly lower levers in induced ESC as compared with that in ES-D3 cells at first, then PKCalpha isoform was found in slightly higher amounts and resumed on 14th day. CONCLUSION: This study was characterized the PKCalpha isoenzyme profile in RA-induced differentiation of ESC into neuron-like cells and suggests that PKCalpha may play a very important role in differentiation of mouse ESC along the neuronal pathway.

Animals↗

PubMed2003

[Confocal microscopy in transplanted human corneas].

PURPOSE: The aim of the study was in-real time observation and morphological evaluation of the transplanted human corneas, using confocal 2.0 microscopy. METHODS: Twelve eyes of 12 patients on 3-7 days after lamellar keratoplasty (LKP), 8 eyes of 8 patients at 1 year after LKP, and 10 eyes of 10 patients at 3-7 days after penetrating keratoplasty (PKP), 11 eyes of 11 patients at 1 year after PKP were examined by confocal 2.0 microscopy in vivo. Images were recorded by continuously focusing the optical section through the full thickness central cornea. RESULTS: 3-7 days after LKP, small stromal cells, cranny-like dark strias and nerves were seen in transplanted corneas. There were highly reflective regions and dots in the lamellar interface. One year after LKP, the nerves disappeared. There were less highly reflective regions, but the dots still can be seen. Some gross dark strias were seen in the posterior stroma and the density of the endothelium cells was normal. 3-7 days after PKP, keratocytes were activated. Nerves and gross dark strias could be seen in transplanted corneas. The density of the endothelium cells was normal, with some highly reflective dots deposited among them. One year after PKP, the nerves disappeared, and strias still existed. The density of the endothelium cells decreased significantly. CONCLUSIONS: Confocal scanning microscopy is a new tool for the study of cellular morphology and construction of transplanted human corneas. It can be used in the evaluation of operation effect, clinical observation and follow-up.

Adult↗

PubMed2002

[Relative quantitative analysis of corneal immunogenicity].

OBJECTIVE: To quantitate the relative immunogenicity of three major corneal cell layers (epithelium, stroma, and endothelium) among the whole corneal immunogenicity respectively. METHODS: Cellular immunity: Three porcine major corneal cell layers were heterotopically allografted to subcutaneous layer of 40 BALB-c mice respectively. Twelve days later, the peripheral white blood cells of recipients were double directly stained with anti-mouse IgG-fluorescein conjugate mAb and analyzed by flow cytometry. Humoral immunity: The suspension of porcine cornea or corneal epithelium was used as antigen to produce anti-porcine immune sera in C57BL-6 mice. With the anti-porcine immune sera, enzyme linked immunosorbent assay was performed. RESULTS: On cellular immunity detection, the immunogenicities of intact endothelium, epithelium and stroma, equal in thickness, were 70.75%, 27.63%, and 1.62% respectively. On humoral immunity detection, the immunogenicities were 62.11%, 31.77% and 6.12% in the three respective layers of equal thickness. CONCLUSION: These findings indicate that the immunogenicity of corneal stroma is the lowest among the three corneal cell layers of equal thickness. The preliminary results provide the experimental basis for clinical application of xenogenic graft of corneal stroma.

Animals↗

PubMed2002

[Corneal topography and thickness in keratoconus].

OBJECTIVE: To evaluate the corneal topography and pachymetry in keratoconus. METHODS: Thirty-five patients (47 eyes) with keratoconus were evaluated by Orbscan corneal topography system. The location, anterior and posterior surface elevation, curvature and pachymetry were analyzed with special reference to the central point of cornea, the apex, the highest point of posterior surface, the point with greatest curvature and the thinnest point. The correlation between the point of greatest curvature and the apex was also analyzed. RESULTS: The mean elevation of the apex was (0.102 +/- 0.044) mm, the mean distance between the apex and the central point of cornea was (0.686 +/- 0.376) mm. Most of the apexes, the points with the highest posterior surface, the points with the greatest curvature were located in the inferior temporal quadrant, followed by inferior nasal, superior temporal and superior nasal quadrant. The SimK of keratocunus was (60.9 +/- 10.4/53.4 +/- 8.7) D and the mean powers within 3 mm and 5 mm were (57.1 +/- 8.0) D and (50.7 +/- 5.4) D respectively. High correlation between apex elevation and the apex curvature was found (r = 0.697, P < 0.01), and there was also negative relationship between apex elevation and corneal thickness (r = -0.563, P < 0.01). CONCLUSIONS: The Orbscan system can provide accurate and subtle information about the shape and thickness of keratoconus, which can help us to learn this disorder deeply, especially in the early diagnosis of this disease.

Adolescent↗