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Purpose: Integrin-linked kinase (ILK) is a serine-threonine kinase associated with anchorage-independent growth and tumorigenic transformation. Previous studies indicate that overexpression of ILK is common among several types of tumors, and it is involved in the regulation of tumor cell survival under stress. In this study, we examined the effects of ILK expression on tumor cellular response to hyperthermia., Experimental Design: We used an adenovirus-mediated approach to overexpress the ILK gene in a prostate cancer cell line and examine its effects on heat stress-induced cell death. Clonogenic survival, as well as apoptosis, was evaluated in cells that overexpress ILK. In addition, the ability to form tumors in vivo was examined in syngeneic hosts. Finally, potential molecular mechanisms of ILK-mediated resistance to heat were examined by determining the status of a variety of signal transduction pathways., Results: ILK overexpression made tumor cells significantly more resistant to the cell-killing effects of hyperthermia. This was correlated at the molecular level with the down-regulation of hyperthermia-induced activation of stress-activated protein kinase/c-Jun-NH(2)-terminal kinase, p38 mitogen-activated protein kinase activities, and caspase 9. The overexpression of ILK was also shown to induce a more rapid tumor growth in a murine prostate cancer cell line, Conclusion: ILK plays an important role in tumor growth and tumor response to hyperthermia treatment.

Objective: To prepare anti-human retinoblastoma (Rb) monoclonal antibody (McAb) by immunization with solid tumor cells and preliminarily study the characteristics of its antigen., Methods: Three Balb/C mice were immunized by intraperitoneal injection of Rb solid tumor cells acquired from enucleation. Spleen lymphocytes were separated from them and fused with myeloma cell line SP2/0. The anti-human Rb McAb was selected by Rb solid tumor cell, SO-Rb(50), SO-Rb(70), etc as antigens. The characteristics of its antigen were preliminarily studied by immunofluorescence, immunohistochemistry and Western blot., Results: After repeatedly cloning them with micro-manipulation equipment, we successfully established 3 hybridoma cell lines secreting anti-human Rb McAb. Of all, 3C6 hybridoma cells could steadily secrete anti-human Rb McAb after they were frozen, resuscitated and passaged repeatedly for 2 years. Its subgroup was IgG(1). Both immunofluorescence and immunohistochemistry examinations demonstrated that the corresponding antigen of McAb 3C6 was specifically and highly expressed in Rb tumor cell membrane and cytoplasm. The other tumor tissues and human normal eye tissues were negative. Western blot analysis preliminarily demonstrated that McAb 3C6 could bind 25 000 protein band of Rb antigen., Conclusions: The anti-human Rb McAb 3C6 established by immunization of solid tumor cells is specifically and highly expressed in Rb tumor cells and has no cross-response with other tumor tissues and normal eyeballs. The antigen molecular weight bounded to this antibody is 25 000 or so, which further shows that it is a possibility for a new unidentified gene to be concerned with the tumor formation of Rb.

Purpose: To clone the variable region gene of light chain of monoclonal antibody against human retinoblastoma and to analyze the characterization of its nucleotide sequence as well as amino acid sequence., Methods: Total RNA was extracted from 3C6 hybridoma cells secreting specific monoclonal antibody (McAb) against human retinoblastoma (RB), then transcripted reversely into cDNA with olig-dT primers. The variable region of the light chain (VL) gene fragments was amplified using polymerase chain reaction(PCR) and further cloned into pGEM -T Easy vector. Then, 3C6 VL cDNA was sequenced by Sanger's method. Homologous analysis was done by NCBI BLAST., Results: The complete nucleotide sequence of 3C6 VL cDNA consisted of 321 bp encoding 107 amino acid residues, containing four workframe regions (FRs) and three complementarity-determining regions(CDRs) as well as the typical structure of two cys residues. The sequence is most homological to a member of the Vk9 gene family, and its chain utilizes the Jkl gene segment., Conclusion: The light chain variable region gene of the McAb against human RB was amplified successfully, which belongs to the Vk9 gene family and utilizes Vk-Jkl gene rearrangement. This study lays a good basis for constructing a recombinant antibody and for making a new targeted therapeutic agents against retinoblastoma.

Objective: To study the incidence of sinusitis in patients with idiopathic orbital inflammatory pseudotumor(IOIP) and the relationship between sinusitis and IOIP., Method: 209 IOIP cases from Jan 1, 1978 to Dec 31, 1999 in Eye Hospital, Zhongshan Ophthalmic Center, Sun Yat Sen University of Medical Sciences were reviewed. Data showed that 36 of 209 IOIP cases suffered from sinusitis including 21 cases of maxillary sinusitis, 15 case of ethmoid sinusitis, 3 case of frontal sinusitis and 3 case of sphenoid sinusitis. The total incidence of sinusitis is 17.2%. Proptosis, restrictive movement of the involved eye, eyelid or conjunctive congestion, decreased eyesight and palpable orbital masses were the main clinical findings. All patients showed a space occupied lesion in their orbits. 27 patients received combined surgical management and corticosteroids treatment. 9 patients received systemic corticosteroids treatment., Result: Therapy showed that the full recovery response rate was 33.3% (12/36) and the partial recovery response was 50.0% (18/36). The recurrence rate was 16.7% (6/36)., Conclusion: Some patients suffered from sinusitis and sinusitis may be a cause in the etiology of IOIP.

Objective: To determine whether polymerase chain reaction (PCR) for the assay of B-cell gene rearrangement in patients with orbital lymphoproliferative disorders could be useful in the diagnosis of lymphoma, especially in differentiating a benign lesion from a malignant one., Methods: In addition to clinical, pathological and immunohistochemical evaluations, 48 cases of orbital lymphoproliferative disorders were examined for immunoglobulin heavy chain (IgH) gene rearrangement and bcl-2/J(H) fuse gene rearrangement by means of PCR to amplify the third frame work region (FR3) and bcl-2/J(H) fuse gene with formalin-fixed and paraffin-embedded tissues., Results: The PCR using primer FR3A showed that 22 cases had discrete single products, which were within the molecular weight range of 100 to 120 bp, and were therefore interpreted as monoclonal. The positive rates of FR3 region of IgH gene rearrangement in patients with malignant lymphoma, benign reactive lymphoid hyperplasia and lymphocytic inflammatory pseudotumor were 75.0% (15/20), 40.0% (4/10) and 16.7% (3/18) respectively. The four patients with simultaneously bilateral ocular adnexal lymphoid neoplasm exhibited identical clonal IgH gene rearrangement patterns. A faint smear pattern or no band at all was demonstrated in the remaining 26 cases, representing polyclonal populations of lymphoid cells. The PCR using bcl-2 primer showed that 6 cases had discrete single products that were within the molecular weight range of 100 to 300 bp, and were interpreted as having bcl-2/J(H) fuse gene. They were 2 cases of follicular B-cell lymphomas, 1 case of diffuse mixed large and small cell B-lymphoma and 3 cases of diffuse small cell B-lymphoma. A faint smear pattern or no band at all was demonstrated in the remaining 17 cases of lymphomas, lymphocytic inflammatory pseudotumor and benign reactive lymphoid hyperplasia. The positive rates of bcl-2/J(H) fuse gene rearrangement in patients with malignant lymphoma, benign reactive lymphoid hyperplasia and lymphocytic inflammatory pseudotumor were 30.0% (6/20), 0% (0/10), and 0% (0/18), respectively., Conclusions: Molecular genetic analysis by PCR using the FR3A primer is helpful in the diagnosis of orbital lymphoproliferative disorders, especially those in which the diagnosis can not be made by routine histopathological finding and immunohistochemistry evaluation. However, the rate of bcl-2/J(H) fuse gene rearrangement is too low to be suitable for clinical diagnosis of orbital lymphoproliferative disorders.

Purpose: The pluripotent embryonic stem cells can differentiate into various kinds of normal tissues. There is no previous report on the differentiation of embryonic stem cell in the intraocular environment. In this paper, the authors tried to investigate the intraocular growth character of mice embryonic stem cells in nude mice., Methods: Murine embryonic stem cells were cultured and maintained in an undifferentiated state in vitro. They were transplanted into the right eyes of 20 nude mice by microinjection under operating microscope. Animal eye observation, light microscope and immunohistochemical examinations were implemented., Results: Two to three days after transplantation, small pieces of gray-white material could be viewed in the vitreous cavity. Between the 15th and 20th day, the gray-white mass grew into the anterior chamber in 4 nude mice eyes. Then, the mass at the anterior chamber extended extraocularly. On the 30th day, a remarkable proptosis was observed in two of the four nude mice. In 6 to 45 days, the mice were executed for morphological examination which showed the following typical structures: (1) Undifferentiated cells with prominent nucleolius. (2) Flexner-Wintersteiner-like rosettes. (3) Medulloepithelioma-like structure: the cells were arranged in sheets, cords, tubes, and cysts. (4) Large, spindle-or astrocyte-like cells. (5) Cartilage-like structure. Immunohistochemically, most of the cells were highly positive in NSE staining and a few cells were moderately positive in GFAP staining., Conclusions: Both animal eye findings and morphologic examinations certificated that the transplanted embryonic stem cells could grow in the eyes of nude mice and differentiate into intraocular medulloepithelioma.