Adenocarcinoma, Mucinous ; enzymology ; pathology ; Cathepsin B ; metabolism ; Cathepsin D ; metabolism ; Colorectal Neoplasms ; enzymology ; pathology ; Humans ; Lymphatic Metastasis ; Neoplasm Invasiveness

Diurnal changes of lysosomes including ultrastructural changes of phagosomes and acid phosphatase reactions in phagosomes, as well as diurnal biochemical changes in cathepsin D activity, were studied in the retinal pigment epithelium (RPE) of the rabbit. The rabbit was maintained on a natural light-dark cycle over seven days in fall and was sacrificed at various times during the day and night. The number of lysosomes or phagosomes in the RPE was the highest at 1.5 hours after exposure to sunlight (8:00 AM), and thereafter decreased with time. Three types of phagosomes were observed and acid phosphatase reactions were different in each type of phagosome; the fresh phagosomes were negative or positive, lamellar bodies positive, and dense bodies partially positive. The biochemical activity of cathepsin D was the highest at 8:00 AM, and this was consistent with the time of peak in phagocytic activity in the RPE. This report shows that phagocytic activity in the RPE occurred in the early stage after exposure to sunlight, and that fresh phagosomes were sequentially degraded to lamellar or dense bodies. Cathepsin D activity also increased, and this was consistent with the phagocytic activity in the RPE.
Acid Phosphatase/metabolism ; Animals ; Cathepsin D/metabolism ; Cell Count ; Choroid/metabolism/ultrastructure ; Circadian Rhythm/*physiology ; Lysosomes/*metabolism/ultrastructure ; Phagosomes/*metabolism/ultrastructure ; Pigment Epithelium of Eye/*metabolism/ultrastructure ; Rabbits

BACKGROUNDHyaluronidase (Hyase) is an enzyme which hydrolyses hyaluronan (HA), a large nonsulfated glycosaminoglycan. Several genes have been identified to code for hyaluronidases in humans. Its role has only recently been underlined in the invasion of prostate cancer, colonic cancer, and breast cancer. Moreover, the findings were in agreement with some experimental results which showed that HA-derived oligosaccharides had angiogenesis-promoting activity. All these findings prompted us to investigate factors that had been characterized as putative invasive factors in different human breast cancer-derived cell lines.

METHODSWe selected two series of human breast cancer-derived cell lines whose expression of estrogen receptors (ER) was previously published. Hyaluronidase secretion in culture medium and expression of matrix metallo-proteinase (MMP)-9, cathepsin-D (cath-D) and vascular endothelial growth factor (VEGF) by cells were determined. We also investigated cell invasiveness in the Matrigel invasion assay, and studied the capability of cancer cells to promote in vitro formation of tubules by endothelial cells.

RESULTSER(-) cells secreted significantly more hyaluronidase (P < 0.001) and expressed significantly more VEGF (P < 0.01), MMP-9 (P < 0.05) and cath-D (P < 0.0001) than ER(+) cells. Invasion through Matrigel by ER(-) Hyase(+) cells was significantly higher than that by ER(+) Hyase(-) cells (P < 0.05). In both cases, invasion was decreased by heparin (P < 0.05). When ECV-304 endothelial cells were co-cultivated in millicell chambers with cancer cells, ECV-304 cells were induced to form tubules. Tubule formation was demonstrated to be more prominent with ER(-) Hyase(+) cells than with ER(+) Hyase(-) cells (P < 0.05).

CONCLUSIONInvasive features of ER(-) breast cancer cells can be characterized in vitro by an invasive Matrigel assay, as the induction of tubule formation by ECV-304 endothelial cells, higher secretion of hyaluronidase, and higher expression of proteinases MMP-9, cath-D, and the angiogenesis promoting factor VEGF.

Breast Neoplasms ; metabolism ; Cathepsin D ; metabolism ; Cell Line, Tumor ; Humans ; Hyaluronoglucosaminidase ; metabolism ; Immunohistochemistry ; Matrix Metalloproteinase 9 ; metabolism ; Neoplasm Invasiveness ; genetics ; Receptors, Estrogen ; genetics ; Vascular Endothelial Growth Factor A ; metabolism

OBJECTIVE: To study the expression of cathepsin-B and -D in different time point after traumatic brain injury. METHODS: Traumatic brain injury (TBI) model was established on rats, cathepsin-B and cathepsin-D immunofluorescence staining and confocal microscope analysis were performed. Positive cells were counted by confocal microscope and image analysis techniques were used to determine the morphological changes in each group. RESULTS: Immunofluorescence staining results showed that cathepsin-B was activated 1 hour after TBI while cathepsin-D was not activated until 12hour after TBI. Both of them got to their peak during 4 to 8days, and kept a high level of activating 32days after TBI. Cathepsin-B and -D positive cells did not merge with caspase-3 positive cells until 6 h after TBI. CONCLUSION Cathepsin-B and -D could be the diagnostic markers of TBI and can estimating time course of lateral TBI. They blocked caspase-3 activation at the beginning period after TBI and started to promote cell death with caspase-3 6 h after TBI.
Animals ; Brain/pathology* ; Brain Injuries/pathology* ; Caspase 3/metabolism* ; Cathepsin B/metabolism* ; Cathepsin D/metabolism* ; Disease Models, Animal ; Forensic Pathology ; Hippocampus/pathology* ; Immunohistochemistry ; Lysosomes ; Male ; Neurons/metabolism* ; Rats ; Rats, Sprague-Dawley ; Time Factors

OBJECTIVETo explore the Zedoary oil on A549 cell line of collagen deposition cat D and cat K expression.

METHODThe A549 cell line were treat by Zedoary oil on four different concentrations (0, 40, 80, 120 mg x L(-1)) in different time. Dynamic changes of collagen in A549 cell using Picric-sirius red method. Cat D and Cat K expression of level were detected by using western blot.

RESULTThe collagen content showed that Zedoary oil had an inhibitory effect on the deposition of A549 cells. The results of western blot showed that the expression of cat D and cat K were up-regulated significangly in A549 cells of Zedoary oil groups compared with that in controls.

CONCLUSIONA549 cell of collagen deposition were reduced by Zedoary oil. The effects may due to the up-regulation of cat D and cat K.

Animals ; Blotting, Western ; Cathepsin D ; metabolism ; Cathepsin K ; metabolism ; Cell Line, Tumor ; Collagen ; metabolism ; Curcuma ; chemistry ; Gene Expression Regulation, Neoplastic ; drug effects ; Plant Oils ; isolation & purification ; pharmacology ; Up-Regulation

OBJECTIVETo establish differential proteomics profiles of glioblastoma cell lines from Chinese, and to provide reference for future basic studies.

METHODSTotal protein was extracted from 3 glioblastoma cell lines, CHG-5, TJ899 and TJ905. After normalization, the total protein was presented by two-dimensional (2D) electrophoresis, scanned and analyzed. Some of the identified protein spots were verified by immunocytochemistry of cell lines and immunohistochemistry of solid tumors. The glia cells were used as the control throughout the study.

RESULTSA total of 13 differential protein spots were selected, and eventually 10 were identified as unique proteins. These 10 proteins were involved in cytoskeleton forming, cellular metabolism, tumor migration, stress and inflammatory reaction. Immunocytochemistry and immunohistochemistry further confirmed these proteins present in the solid tumors.

CONCLUSIONDistinct differential proteomics profiles exist in glioblastoma cell lines and normal glia cells, likely related to the transformation of normal glia to glioma.

Brain Neoplasms ; genetics ; metabolism ; Cathepsin D ; metabolism ; Cell Line, Tumor ; Gene Expression Profiling ; Glial Fibrillary Acidic Protein ; metabolism ; Glioblastoma ; genetics ; metabolism ; Humans ; Microfilament Proteins ; metabolism ; Neuroglia ; metabolism ; Proteomics ; methods

OBJECTIVE: To study the function of lymphangiogenesis and the expression of Cathepsin D (Cath-D) in laryngeal carcinoma metabasis and clinical pathology character. METHOD: The expression of Cath-D were detected in 76 laryngeal carcinoma with immunohistochemistry (SP method). Podoplanin was used as the marker of lympgatic vessel endotheliocytes to label lympgatic vessel in 76 laryngeal carcinoma,lymphatic microvessel density were measured,and the paraneoplastic tissues was used as control group. RESULT: The positive rate of Cath-D in paraneoplastic tissue, laryngeal carcinoma and in pathology classification, in clinical stage, in cervicale lymphonode metastasis negative and positive group were significantly different. However, there had no difference between the positive rate of Cath-D in the age specific and clinical classification. c) The lymphatic microvessel density in paraneoplastic tissue, laryngeal carcinoma and clinical stage, in glottic carcinoma and supraglottic carcinoma, in cervical lymphonode metastasis negative and positive group were significantly different; but there had no difference in age-specific and pathology classification. CONCLUSION (1) The high expression of lymphatic microvessel density and the increasing expression of Cath-D could promote cervical lymphonode metastasis in aryngeal carcinoma. (2) There had a correlation between the high expression of lymphangiogenesis and Cath-D in laryngeal carcinoma, and had cooperation in aryngeal carcinoma lymphonode metastasis.
Biomarkers, Tumor ; metabolism ; Carcinoma ; metabolism ; pathology ; physiopathology ; Cathepsin D ; metabolism ; Female ; Glottis ; Humans ; Immunohistochemistry ; Laryngeal Neoplasms ; metabolism ; pathology ; physiopathology ; Lymphangiogenesis ; physiology ; Lymphatic Metastasis ; Lymphatic Vessels ; metabolism ; Male ; Membrane Glycoproteins ; metabolism

The purpose of study was to explore the possible functions of Bcl-xL in the glucosamine sulfate-induced apoptosis of chronic myeloid leukemia K562 cells. Light microscopy and Wright-Giemsa staining were used to investigate the morphologic evidences for apoptosis of K562 cells induced by glucosamine sulfate (GS); immunofluorescence was used to observe the translocation of cathepsin D and cytochrome C during the apoptosis; Western blot was performed to detect the expression of Bcl-xL, Bid, Bax in K562 cells treated by GS. The results showed that many vacuoles were observed in the cytoplasma of the K562 cells treated by GS; fluorescent signals of cathepsin D and cytochrome were fransformed from granules to disperse form by using immunofluorescence; the expression of Bcl-xL was found down-regulated in K562 cells treated by GS, but not in the cells pre-treated with pepstatin A; the significant changes were not detected in expression of Bax and Bid protein before or after apoptosis. It is concluded that Bcl-xL protein may mediate relationship between cathepsin D and mitochondia pathway, Cathepsin D may play an important role in the GS inducing apoptosis of K562 cells through downregulation of Bcl-xL expression.
Apoptosis ; drug effects ; physiology ; BH3 Interacting Domain Death Agonist Protein ; metabolism ; Blotting, Western ; Cathepsin D ; metabolism ; Cytochromes c ; metabolism ; Fluorescent Antibody Technique ; Glucosamine ; pharmacology ; Humans ; K562 Cells ; bcl-2-Associated X Protein ; metabolism ; bcl-X Protein ; metabolism ; physiology

OBJECTIVEAutophagy is involved in several neurodegenerative diseases and recently its role in acute brain injury has won increasing interest. Spinal cord injuries (SCIs) often lead to permanent neurological deficit. Therefore, in this study, we examined the pro?les of autophagy-linked proteins (MAP-LC3) after SCI to investigate whether the expression of autophagy contributes to neurological deficit after SCI.

METHODSAdult female Sprague-Dawley rats were used and randomly divided into control and SCI groups. All the rates received laminectomy at T8-T10 level. Those in the SCI group received additional exposure of the dorsal surface of the spinal cord, followed by a weight- drop injury. Thereafter we investigated the expression levels of MAP-LC3, beclin-1, Cathepsin D and the beclin-1-binding protein bcl-2 by western blot analysis at 12 h, 24 h, 3 d, 7 d, 21 d and 28 d. One-way ANOVA with Tukey post hoc test was used to compare data between groups.

RESULTSWe observed significant increase in the level of LC3 (LC3-II/LC3-I) at 3 d and 7 d after SCI when compared with the sham group. While the level of beclin-1 and ratio of beclin-1/bcl-2 was found to have increased from 12 h to 24 h after injury. Cathepsin D expression was also elevated at 7 d (P<0.01).

CONCLUSIONBased on the above mentioned data, we proposed that autophagy plays a role in the manifestation of cell injury following SCI.

Adaptor Proteins, Signal Transducing ; metabolism ; Animals ; Apoptosis Regulatory Proteins ; metabolism ; Autophagy ; physiology ; Beclin-1 ; Blotting, Western ; Cathepsin D ; metabolism ; Disease Models, Animal ; Female ; Laminectomy ; Microtubule-Associated Proteins ; metabolism ; Rats ; Rats, Sprague-Dawley ; Spinal Cord Injuries ; metabolism

Cathepisn D plays a key role in early process of apoptosis before mitochondrion damage and caspases activations, and also involves in Alzheimer's disease (AD). Glycosaminoglycans (GAGs) have been suggested to inhibit the progress of apoptosis. Fucoidan, a nature GAGs mimetic, is shown as a potential candidate for neuroregressive disease. Here we reported PC12 cells response to oxidative stress with clear cathepsin D release, followed by caspase-3 activation. We found that fucoidan treatment can alleviate cathepsin D and caspase-3 activation, and improve cell survival. Furthermore, for the first time, fucoidan was shown to directly inhibit human liver cathepsin D by a dose-dependent way. These results support that cathepsin D involves in early apoptosis, suggest that fucoidan can decrease apoptosis at lysosome-cathepsin D level, which opens a new therapeutic approach to AD.
Alzheimer Disease ; drug therapy ; metabolism ; Animals ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Caspase Inhibitors ; Cathepsin D ; antagonists & inhibitors ; metabolism ; Cell Line, Tumor ; Cell Survival ; drug effects ; Humans ; Hydrogen Peroxide ; metabolism ; Oxidative Stress ; drug effects ; PC12 Cells ; Polysaccharides ; pharmacology ; Rats