Apoptosis-induced Proliferation in UV-Irradiated Human Conjunctival Epithelial Cells

A pterygium is a benign growth that develops on the conjunctiva and, in some cases, extends to the cornea and interferes with vision. Excessive exposure to ultraviolet (UV) light is one of the causes of pterygium development. We previously reported that UV-induced apoptosis is led by production of reactive oxygen species (ROS) that activate p38 mitogen-activated protein kinase (MAPK) in human conjunctival epithelial (HCE) cells. Also, ROS-dependent induction of interleukin-11 (IL-11) has been reported to upregulate MAPK pathways, which results in compensatory proliferation. In this study, we examined the effect of UV exposure on HCE cells, in terms of change in apoptosis, ROS generation, phosphorylation of c-Jun N-terminal kinase (JNK), levels of IL-11 (a key cytokine in tissue repair and compensatory proliferation), production of activator protein 1 (AP-1), and expression of c-myc, c-fos and c-jun (which provides evidence of healthy cell proliferation). Apoptosis in HCE cells was induced by UV light irradiation (312nm, 4.94mW/cm2). Apoptosis was measured using the Muse Annexin V and Dead Cell Assay Kit. ROS generation was measured by using 5-(and 6-) chloromethyl-2\u277\u27-dichlorodihydrofluorescein diacetate, acetyl ester. JNK phosphorylation, IL-11 levels and AP-1 production were measured by enzyme-linked immunosorbent assays (ELISAs). Imnunocytochemical staining was used to measure c-myc, c-fos and c-jun expression. UV irradiation increased ROS generation, phosphorylation of JNK, and apoptotic cell count. IL-11 levels and AP-1 production were significantly increased by UV irradiation. The irradiated cells had increased expression of c-myc, c-fos and c-jun, and treatment of the cells with IL-11 significantly increased expression of c-myc, c-fos and c-jun. These results suggest that the release of IL-11 from UV-induced apoptotic HCE cells and surrounding healthy cells could promote proliferation to maintain homeostasis

The Effectiveness of Combined Medical Therapy and Hemodialysis for Hypercalcemia in Anaplastic Lymphoma Kinase-negative Anaplastic Large Cell Lymphoma

A 71-year-old man with a right lower abdominal quadrant epithelial tumor developed gradually worsening lumbago and dysbasia. He became comatose and was admitted to our hospital. He had swelling of the left axillary lymph nodes and necrosis of the 4.0-cm diameter abdominal tumor, which infiltrated the subcutaneous tissues. He was hypercalcemic (16.7mg/dl), and had elevated levels of soluble interleukin-2 receptor (24,090U/ml) and parathyroid hormone-related protein (5.4pmol/l). Computerized tomography (CT) showed left axillary lymphadenopathy, splenomegaly, and a right abdominal-wall mass that was described as anaplastic large cell lymphoma upon pathology. Brain radiography and CT revealed multiple lesions infiltrating the cranium. Magnetic resonance imaging showed diffuse low signal intensity throughout the vertebral spine. The patient was diagnosed with anaplastic lymphoma kinase (ALK) -negative anaplastic large cell lymphoma with hypercalcemia. Fluid replacement and drug therapies including calcitonin had no effect on the hypercalcemia or the coma. The patient\u27s serum calcium concentration decreased after hemodialysis (calcium dialysate concentration, 5mg/dl) and subsequent zoledronic acid hydrate therapy. His consciousness improved by the fifth day of treatment. This rare case of hypercalcemia in ALK-negative anaplastic large cell lymphoma improved with combined medical and hemodialysis therapy

Vildagliptin Improves Glucose Tolerance and Decreases Plasma Triglycerides in Sprague-Dawley Rats

The number of patients with lifestyle-related diseases, including type 2 diabetes, is increasing. The onset of type 2 diabetes can be prevented by dietary and exercise interventions, as well as drug therapy. Dipeptidyl peptidase-4 inhibitors and glucagon-like peptide-1 receptor agonists have attracted attention recently as treatments for diabetes, and incretin hormones have been reported to have a protective effect on pancreatic β-cells. It is not clear whether vildagliptin (VIL) can prevent the progression of lifestyle-related disease. Thus, in the present study, Sprague-Dawley rats were fed a high-fat diet with sucrose water (HFDS) to determine whether VIL could inhibit deterioration in glucose tolerance and improve other biomarkers of lipid disorder. Four-month-old male Sprague-Dawley rats were divided into three groups (n = 7 in each group); one group was fed a normal diet for 4 months, whereas the remaining two groups were fed the HFDS, with or without VIL for 4 months. When rats were 7 months of age, they were subjected to an intraperitoneal glucose tolerance test (IPGTT); biomarkers of lipid disorder were measured in 8-month-old rats. There was a decrease in the glucose spike in the IPGTT 10min after loading in the HFDS + VIL group and plasma triglyceride (TG) levels were significantly lower in these rats compared with the HFDS group. The decreased TG levels in HFDS + VIL rats were accompanied by decreases in plasma chylomicron levels. These results suggest that VIL can prophylactically inhibit decreases in pancreatic β-cell function in type 2 diabetes and reduce the risk of cardiovascular disease due to high TG levels. Thus, VIL administration may contribute to the prevention of lifestyle-related disease

Suppressive Effects of Catechins in UV-Induced Cytotoxicity of Human Corneal Epithelial Cells

Photokeratitis is a disease in which the ocular surface is directly affected by oxidative stress caused by exposure to ultraviolet （UV） light and oxygen. It is speculated that the production of free radicals and reactive oxygen species （ROS） is caused by UV-induced cytotoxicity. Recent studies have reported that catechins have antioxidant, antiallergic, antitumor, and antibacterial effects. The aim of our study was to investigate the mechanism of UV-induced cytotoxicity in cultured human corneal epithelial （HCE-T） cells and evaluate the protective effects of the catechins, （?）-epigallocatechin gallate （EGCG） and （?）-epigallocatechin 3-O-（3-O-methyl） gallate （EGCG3”Me）, on apoptosis. HCE-T cells were UV irradiated at 312nm （4.94mW/cm2, 296mJ/cm2）. EGCG and EGCG3”Me were dissolved in methanol and adjusted to 5, 10, or 20?M. Absorption was measured from 250 to 400nm. EGCG and EGCG3”Me were pre-incubated for 1 hr. After UV irradiation, membrane lipid peroxide, tumor necrosis factor （TNF）-α production, ROS generation, caspase-3 and -8 activities, mitochondrial membrane potential, and cytochrome c levels were measured. Both EGCG and EGCG3”Me had UV absorption, and increased with concentration dependently. The increases in the levels of membrane lipid hyperoxidation, activation of caspase-3 and -8, production of TNF-α and ROS were found, by UV irradiation, to be significant. But these levels were significantly decreased by pretreatment with EGCG and EGCG3”Me. There were no changes in mitochondrial membrane potential and cytoplasmic cytochrome c levels after UV irradiation. Oxidative stress occurs early near the cell membrane in response to UV irradiation. As a result, TNF-α is induced, leading to apoptosis mainly through caspase-8 activation. Conversely, EGCG and EGCG3”Me absorb UV light directly and inhibit lipid peroxidation in the cell membrane. Catechins inhibit the apoptosis cascade by inactivating caspase-3 and caspase-8

Construction of a Novel Single Double-Conditional shRNA Expression Vector

Silencing of gene expression by RNA interference (RNAi) has become a widely used tool for assessing gene function in a fast and easy manner. An important advance in the RNAi field was the discovery that plasmid-based short hairpin RNA transcription can substitute for synthetic small interference RNAs both in vitro and in vivo. The constitutive knockdown of gene expression by RNAi can limit the scope of experiments, especially if the inhibition of genes leads to cell lethality, which prevents in vivo functional analysis. We have generated double-conditional short hairpin RNA from a single vector that can regulate its own transcription by a combination of tetracycline-inducible and Cre-loxP systems. This vector will be useful for the control of short hairpin RNA expression in a spatially-regulated manner by Cre recombinase as well as a temporally-regulated manner by tetracycline induction. These control features provide for the specific silencing of genes to facilitate functional genetic analysis in mammals in vivo