Objective To investigate the effect of hypoxia on expressions of erythropoietin(EPO)mRNA and protein in retinal M?ller cells cultured in vitro. Methods Retina tissues from the new-born Wistar rats were dissected into cell suspension after digested by pancreatin.M?ller cells were separated and purified by mechanical concussion and blowing and striking method.The expression of EPO mRNA and protein under the condition of hypoxia was detected by semi-quantitative reverse transcriptase(RT)-polymerase chain reaction(PCR)and immunocytochemical method. Results Retinal M?ller cells were cultured successfully,95% of which were positively stained by glial fibrillary acidic protein(GFAP).Positively stained EPO protein was located in the cytoplasm and protuberance.The expression of EPO mRNA and protein was faint in the normal retinal M?ller cells,but increased obviously and time-dependently after hypoxia. Conclusion Expression of EPO mRNA and protein increases in M?ller cells after hypoxia,which may be one of the protective factors for the nerves in anoxic retinopathy.

OBJECTIVE: To investigate the effects of Radix notoginseng extracts drug-containing serum on the expressions of apoptosis-regulating proteins including Bax, bcl-2 and p21WAF1 in precancerous gastric cells. METHODS: The N-methyl-N'-nitro-N-nitroso-guanidine (MNNG) transformed eternalized human gastric mucosa epithelium GES-1 cell line (MC cell) was used in vitro as a model of gastric precancerous lesion. The medicated canine serum was prepared by feeding to the adult Beagle dog with Radix notoginseng extracts and obtaining the serum after 2-hour medication. MC cells were cultured with medicated canine serum (medicated serum group) or non-medicated canine serum (normal control group) for 72 hours. Expressions of Bax, bcl-2 and p21WAF1 proteins were detected by immunocytochemical assay and the average optical density of the cells was determined by an image analysis system. RESULTS: Compared with those of the normal control group, Bax and p21WAF1 expressions in medicated serum group were significantly enhanced (P<0.01), while the expression of bcl-2 was significantly reduced (P<0.01). CONCLUSION: Radix notoginseng extracts may inhibit the proliferation and promote the apoptosis of precancerous gastric cells through altering expressions of the bcl-2, Bax and p21WAF1 genes.

OBJECTIVE: To study the apoptosis-promoting effect of the serum from Panax notoginseng extracts-fed dog on precancerous gastric cells by means of flow cytometry. METHODS: In the experiment, we adopted eternalized human gastric mucosa epithelium GES-1 cells transformed by N-methyl-N'-nitro-N-nitroso-guanidine (MNNG) (MC cells) as the model of precancerous lesions for study in vitro. We took the serum of a dog before and at two different points of time (2 and 6 hours) after feeding the dog with Panax notoginseng extracts for experiment. The MC cells were cultured in mediums with different concentrations of the medicated serum at 2- or 6-hour point of time for 72 hours. By means of flow cytometry, we examined the apoptosis-promoting effects of the serums on the MC cells. RESULTS: The medicated serums at these 2 points of time had significant effects in promoting MC cell apoptosis. The proportions of apoptotic cells in culture mediums with medicated serums had a significant increase as compared with those in culture mediums with non-medicated serums (serum obtained before administration of extracts to the dog) under the same conditions (P<0.05). The number of MC cells in G(0)/G(1)phase was decreased (P<0.05) and that in G(2)/M phase increased (P<0.05), while no consistent changes were observed in S phase. CONCLUSION: The medicated serums obtained at the two different points of time have significant apoptosis-promoting effects on MC cells. They decrease the number of MC cells in G(0)/G(1) phase and increase the number of MC cells in G(2)/M phase. This is probably responsible for the effects of Panax notoginseng extracts in inhibiting the proliferation of MC cells and promoting its apoptosis.

OBJECTIVE: Through cell cultivation, we studied the inhibiting effects of the serum of the dog fed with Panax notoginseng extracts on precancerous gastric cells, trying to find the best time points or periods when the extracts' function was the strongest after administration of the extracts to the dog. METHODS: The experiments adopted eternalized human gastric mucosa epithelium GES-1 cells and MC cells gained from GES-1 cells transformed by N-methyl-N'-nitro-N-nitroso-guanidine (MNNG) as the model of precancerous lesions for study in vitro. We took the serum of a dog before and at different points of time after feeding the dog with Panax notoginseng extracts for experiment. By means of MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay, we examined the inhibiting effects of the serum after culturing the GES-1 and MC cells for 72 hours with different concentration (8%, 4%, 2%) of medicated serum obtained from the dog at different points of time, so as to find that, at which points of time the medicated serum obtained, it would be the most effective. RESULTS: The results showed that the GES-1 and MC cells inhibition rates of medicated serum from the points of 2-hour and 6-hour were the highest, and the culture medium containing 8% of medicated serum from these two points had prominent inhibiting effects on both kinds of cells. The GES-1 cells inhibition rate in culture medium containing 8% of medicated serum from the point of 2-hour was 70.8% (P<0.01) and that of the MC cells was 45.3% (P<0.01). The GES-1 cells inhibition rate in culture medium containing 8% of medicated serum from the point of 6-hour was 88.5%(P<0.01) and that of the MC cells was 42.4% (P<0.01). CONCLUSION: The points of time with the strongest inhibiting effects are 2 hours and 6 hours after being fed with Panax notoginseng extracts. At these two points, the serum is most effective in inhibiting the proliferation of GES-1 and MC cells.

AIM: To investigate the effects of salidroside on intracellular free calcium concentration (([Ca~(2+)]i)), apoptosis, mitochondrial membrane potential (MMP) and activity during injury induced by hypoxia/hypoglycemia in cultured SH-SY5Y cells. METHODS: Mitochondrial activity was measured by methylthiazolyl tetrazolium test. MMP, [Ca~(2+)]i and apoptosis were measured by flow cytometry. RESULTS: SH-SY5Y cells were cultured in a hypoxia/hypoglycemia condition for 2, 4, 6 and 12 h, [Ca~(2+)]i and apoptosis rate significantly increased compared with control group (P

AIM: To investigate proliferation and apoptosis of cultured endothelial cell (ECV-304 cell line) induced by varied concentrations of oxidized low density lipoprotein (ox-LDL). METHODS: Cell morphology, Typan blue test, MTT test, LDH release test, flow cytometry and micro-molecular weight DNA fragment gel electrophoresis of apoptosis were used for the detection of the cytotoxic effects of ox-LDL on ECV-304 cell line. RESULTS: 0 1, 1, 10 mg/L ox-LDL could promote proliferation of ECV-304 cells. When the concentration of ox-LDL reached up to 100 mg/L and above, the distinct cytotoxic effect appeared. Further study showed that the apoptosis rate of endothelial cells, induced by ox-LDL of 150 mg/L and 200 mg/L for 12 hours, are 15 86% and 21 89%, respectively. 18 h and above hours after incubation, the apoptosis rate began to decrease and rate of necrosis increased. CONCLUSION: ox-LDL has strong cytotoxic effects on endothelial cells and could give rise to different pathologic process, such as proliferation, apoptosis prophase, apoptosis and necrosis.

Objective: To observe the expression of cell adhesion molecules ICAM-1 and VCAM-1of cultured rat brain microvascular endothelial cells(MVEC),expecting to explore the mechanisms of new QingKaiLing injection protecting brain from injury of inflammatory cascade in cerebral ischemia diseases.Methods: Rat cerebral MVEC were extracted by separating microvessel sections and collagenase enzymatic digesting,an in vitro ischemia reperfusion model was established(Kreb,95%N2+5%CO2),the protein and mRNA expression of ICAM-1 and VCAM-1 were detected by using immunocytochemical stain and RT-PCR method.Results:The expression of adhesion molecules of model group were significantly higher than those of noral group(P

OBJECTIVETo investigate the expression of cluster of differentiation 44 variant 6 (CD(44V6)) and proliferating cell nuclear antigen (PCNA) in ocular squamous cell carcinomas.

METHODSStreptavidin-biotin complex (SABC) immunohistochemistry was used to explore the expression of CD(44V6) and PCNA in 35 cases of ocular squamous cell carcinomas, 20 cases of papillomas, and 11 cases of normal eyelid tissue.

RESULTSThe CD(44V6) positive rate was 62.9% (22/35) in ocular squamous cell carcinomas, 15.0% (3/20) in papillomas, but not detectable in the 11 cases of normal eyelid tissue. The positive expression rates of CD(44V6) in ocular squamous cell carcinomas were significantly higher than in benign tumors (chi(2) = 11.57, P < 0.01) or control tissue (P = 0.001), and the positive expression rates of CD(44V6) in metastasis were significantly higher than without metastasis (P = 0.049). PCNA labeling indexes (PI) in tumors with CD(44V6) expression were significantly higher than those without (t = 20.21, P < 0.01).

CONCLUSIONSOverexpression of CD(44V6) is correlated with the progress and metastasis of ocular squamous cell carcinomas. CD(44V6) protein positive staining is associated with high PI. CD(44V6) and PCNA are useful for evaluating prognosis.

Carcinoma, Squamous Cell ; chemistry ; pathology ; Eye Neoplasms ; chemistry ; pathology ; Glycoproteins ; analysis ; Humans ; Hyaluronan Receptors ; analysis ; Immunohistochemistry ; Lymphatic Metastasis ; Proliferating Cell Nuclear Antigen ; analysis ; Skin ; chemistry

OBJECTIVETo investigate the relationship between P16 gene (the tumor suppressor gene) and the bcl-2 gene (the apoptosis inhibitor gene) and the incidence and development of malignant eyelid tumors.

METHODSThe streptavidin-biotin-peroxidase complex immunohistochemistry method was used to study the expression of P16 gene and the bcl-2 gene in 96 cases of malignant eyelid tumors.

RESULTSAmong the 96 cases, there were 40 basal cell carcinomas (BCCs), 33 squamous carcinomas and 23 sebaceous carcinoma, with P16 protein positive (nuclear staining) rates 70%, 54.6% and 56.5%, respectively. The P16 positive rate was negatively correlated with the degree of tumor histological differentiation, and the rate difference between the high differentiated carcinomas was significant (P < 0.05). Positive Bcl-2 protein expression was detected in the cytoplasm. All 40 BCC cases were Bcl-2 positive, and nearly all of the tumor cells showed positive cytoplasmic expression, while in the 33 squamous cell carcinoma cases only one showed positive focal reaction, and the staining in the other 32 cases was relatively faint. None of the 23 sebaceous carcinomas expressed Bcl-2.

CONCLUSIONSThe expression of the P16 protein was related to the occurrence and degree of differentiation of malignant eyelid tumors. The overexpression of the Bcl-2 protein suggests that suppression of apoptosis might play a role in the tumorigenesis of BCC.

Adult ; Aged ; Aged, 80 and over ; Carcinoma, Basal Cell ; chemistry ; Carcinoma, Squamous Cell ; chemistry ; Cell Differentiation ; Cyclin-Dependent Kinase Inhibitor p16 ; analysis ; Eyelid Neoplasms ; chemistry ; pathology ; Female ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Proto-Oncogene Proteins c-bcl-2 ; analysis