Distribution of Interleukin-22–secreting Immune Cells in Conjunctival Associated Lymphoid Tissue

PURPOSE: Interleukin (IL)-22 is a cytokine involved in epithelial cell regeneration. Currently, no research studies have analyzed the distribution of the three distinct IL-22–secreting cell populations in human or mouse conjunctiva. This study investigated the distribution of the three main populations of IL-22–secreting immune cells, αβ Th cells, γδ T cells, or innate cells (innate lymphoid cells [ILCs] or natural killer cells), in conjunctival associated lymphoid tissues (CALTs) in human and mouse models. METHODS: We collected discarded cadaveric bulbar conjunctival tissue specimens after preservation of the corneo-limbal tissue for keratoplasty from four enucleated eyes of the domestic donor. The bulbar conjunctiva tissue, including the cornea from normal (n = 27) or abraded (n = 4) B6 mice, were excised and pooled in RPMI 1640 media. After the lymphoid cells were gated in forward and side scattering, the αβ Th cells, γδ T cells, or innate lymphoid cells were positively or negatively gated using anti-CD3, anti-γδ TCR, and anti–IL-22 antibodies, with a FACSCanto flow cytometer. RESULTS: In normal human conjunctiva, the percentage and number of cells were highest in αβ Th cells, followed by γδ T cells and CD3–γδ TCR – IL-22+ innate cells (presumed ILCs, pILCs) (Kruskal-Wallis test, p = 0.012). In normal mice keratoconjunctiva, the percentage and total number were highest in γδ T cells, followed by αβ Th cells and pILCs (Kruskal-Wallis test, p = 0.0004); in corneal abraded mice, the population of αβ Th cells and pILCs tended to increase. CONCLUSIONS: This study suggests that three distinctive populations of IL-22–secreting immune cells are present in CALTs of both humans and mice, and the proportions of IL-22+αβ Th cells, γδ T cells, and pILCs in CALTs in humans might be differently distributed from those in normal mice.

Ocular Surface Squamous Cell Carcinoma with Intraorbital Extension in a Patient with Long-Term Immunosuppression

PURPOSE: To report a case of ocular surface squamous cell carcinoma with intraorbital extension in a patient with renal transplantation and long-term immunosuppressive therapy. CASE SUMMARY: A 59-year-old Korean male presented with a whitish mass in the medial limbus and conjunctiva of the right eye. The patient had undergone renal transplantation 17 years prior due to lupus nephritis and was on systemic immunosuppression with daily prednisolone (10 mg), tacrolimus (5 mg), and mycophenolate sodium (720 mg). The complete excision of the mass was performed and mitomycin C application and amniotic membrane transplantation on the excised area were combined. Histopathological examination revealed the mass was squamous cell carcinoma. There were no abnormal findings on the orbit computed tomography (CT). The patient was additionally treated with topical interferon alpha 2b 6 months postoperatively. One year later, a mass recurred at the same site in the right eye. The complete excision of the mass, mitomycin C application, cryotherapy, and amniotic membrane transplantation were performed. Orbit CT showed a 1.9 cm-sized intraorbital mass involving the medial rectus of the right eye. The orbital exenteration was performed and the intraorbital mass was histologically proven to be squamous cell carcinoma. CONCLUSIONS: Ocular surface squamous neoplasia in patients with renal transplantation and long-term immunosuppressive therapy should be monitored closely for the possibility of orbital invasion.

Transcriptional Analysis of Nod-Like Receptors in a Mouse Model of Experimental Autoimmune Uveitis

PURPOSE: To evaluate the transcription pattern of Nod-like receptors (NLRs), the intracellular sensors, to detect danger signals in murine eyes with experimental autoimmune uveitis (EAU). METHODS: EAU was induced in B6 (C57BL/6) mice by subcutaneous injection of human interphotoreceptor retinoid binding protein and intraperitoneal injection of pertussis toxin. At 1, 2, and 3 weeks post-immunization, the eyeballs were extracted and subjected to histological and molecular assays using real-time reverse transcription polymerase chain reaction. RESULTS: The levels of nucleotide-binding oligomerization domain, Leucine rich Repeat and Pyrin domain 1 (NLRP1), NLRP3, nucleotide-binding oligomerization domain-containing protein 1 (NOD1), and NOD2 transcripts were increased at 2 weeks and gradually reduced thereafter. Notably, NLRP3 showed the highest expression in the eyes with EAU. Similarly, the transcript level of pro-inflammatory cytokine, interleukin-1beta, increased and reached a peak at 2 weeks post-immunization. The retinal structure was severely damaged by inflammation at 3 weeks post-immunization. CONCLUSIONS: Among NLRs, NLRP3 may induce inflammation in eyes after EAU immunization.

Effect of Electrocauterization on the Inflammation of the Conjunctiva in Experimental Animal Model

PURPOSE: Recently, conjunctivochalasis repair surgery using electrocauterization has been gaining popularity. However, patients with electrocauterized conjunctivoplasty tend to complain of more postoperative pain than patients undergoing simple excision with suturing. Therefore, we investigated the effects of electrocauterization on inflammation of the conjunctiva using an experimental animal model and compared these with the effects of simple excision with suturing. METHODS: Ten New Zealand white rabbits underwent cauterization in the right eyes and excision and suturing in the left eyes. For each eye, we excised or electrocauterized the inferior bulbar conjunctiva, 1 mm in width and 6 mm in length, 2 mm from the limbus. A fine-needle electrode was inserted subconjunctivally, and electrocauterization was performed. In the contralateral eye, the corresponding area was excised and re-approximated with 10-0 nylon sutures. Sutures were removed after 14 days. Tissue samples were obtained at 21 days post-procedure, and inflammatory cells were counted in five randomly selected fields (x200) on hematoxylin-eosin stained slides. Tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta concentrations in tears were measured using enzyme linked immunosorbent assays. RESULTS: All cauterized eyes demonstrated smooth surface healing without scarring after 5 days, whereas sutured eyes presented with mild edema with some scarring until the suture was removed. The number of inflammatory cells was significantly greater in sutured eyes compared with cauterized eyes (p = 0.035, Mann-Whitney U-test) at 21 days post-procedure. Tear TNF-alpha and IL-1beta concentrations at 21 days were similar in both groups. CONCLUSIONS: Electrocauterization for conjunctivoplasty seems to be advantageous in terms of inflammation compared with simple suturing and excision.

Effect of ROCK Inhibitor on the Expansion and Wound Healing of Human Corneal Endothelial Cell

PURPOSE: To investigate the effect of ROCK inhibitor Y27632 on the human corneal endothelial cell proliferation in vitro and in vivo. METHODS: Using corneal endothelial cells isolated and cultured from human donor cornea, we compared the effect of Y27632 (10 microM) on the proliferation in vitro by flow cytometry analysis. For the evaluation of the effect of Y27632 (10 mM) in vivo, corneal thickness and wound area were analyzed for the corneal endothelial wound rabbit model induced by transcorneal freezing. RESULTS: Ki67 positive cells were increased in the Y27632 group (9.1 +/- 4.1%) than the control group (8.0 +/- 5.9%), whereas annexin V positive cells in the Y27632 group (2.9 +/- 1.0%) were decreased compared to the control group (4.2 +/- 2.2%). However these were not statistically significant. Wound area after Y27632 application in animal model is concerned, the control group showed significant smaller area (45.6 +/- 0.6 mm2) compared to the Y27632 group (49.3 +/- 0.8 mm2; p = 0.029, Mann-Whitney U test), however, these were not significantly different from the baseline. Corneal thickness was not different between the two groups. CONCLUSIONS: Different from other reports for the effect of Y27632, no significant effect on the proliferation in vitro and wound healing in vivo, regarding human corneal endothelial cell, were found in this study.

Clinical Usefulness of a PET Scan in the Diagnosis of a Recurrent Colorectal Carcinoma

PURPOSE: The aim of this study was to evaluate the potential efficacy of positron emission tomography using 2- [18F]-fluoro-2-deoxy-D-glucose in detecting recurrent colorectal carcinomas. METHODS: Seventy patients suspected of having recurrent colon cancer were evaluated using PET from January 2001 to March 2003. The PET results were compared with those of computed tomography and clinical examination over 6 months. RESULTS: Among the 70 patients, 17 patients had abnormal CEA levels and no abnormal findings with conventional radiologic methods (group 1), 29 had equivocal findings on computed tomography and other radiologic studies (group 2), and 24 were proven to have recurrent colorectal carcinoma (group 3) respectively. The sensitivity and the specificity of the PET scan, which were proved to be higher then those of CT (93% vs. 72.2%, 88% vs. 48.5%), were 85.7%, and 80%, respectively, in group 1, 86.7%, and 85.7% in group 2 and both 100% in group 3. The interval between diagnosis using PET and conventional studies was 1.9 months. CONCLUSIONS: Positron emission tomography is more sensitive than computed tomography in detecting recurrent colorectal carcinomas. When conventional studies show no abnormal findings except for CEA levels, PET can be a valuable imaging tool in detecting recurrent colorectal cancer and can prevent other unnecessary exploratory procedures.

Clinical Evaluation of Tc-99m Ciprofloxacin for the Diagnosis of Infectious Disease / 감염

BACKGROUND: For the evaluation of infectious or inflammatory disease, Tc-99m HMPAO WBC scan has been used. But techniques involving radiolabelled leukocytes scan were difficult, and have failed to distinguish between bacterial infection and non-bacterial inflammation. To overcome this difficulty, we evaluated Tc-99m Ciprofloxacin in clinically suspected bacterial infection. METHODS: We used this agent for 31 patients who had suspected sites of infection. And then we compared the imaging results of these patients with those of radiolabelled leukocyte scan. RESULTS: There was no adverse effect. 29 patients was involved for the interpretation. Infective spondylitis was the most common suspected disease (41%). Bateriologic culture was positive in 10 patients. The most common organism was meticillin-susceptible S. aureus (5 cases). The sensitivity and specificity of Tc- 99m Ciprofloxacin was 83.3% and 66.6%, and those of Tc-99m HMPAO WBC was 62.5% and 83.3%, repectively. For the diagnosis of infective spondylitis the sensitivity of Tc-99m Ciprofloxacin was 80.0% and that of Tc-99m HMPAO WBC was 40.0%. CONCLUSOIN: Tc-99m Ciprofloxacin shows relatively high sensitivity and specificity. Tc-99m Ciprofloxacin was superior to Tc-99m HMPAO WBC for the diagnosis of infection in axial skeleton.