Objective To investigate the different killing effect to human pulmonary adenocarcinoma cell line cells GLC-82 with coexpressed double suicide genes compared with single gene. Methods Recombinant expression vectors containing CD (cytosine deaminase) and/or TK(thymidine kinase) gene under CMV promoter were constructed successfully. The vectors were transfected to GLC-82 tumor cell lines by use of lipofectamine. The clones were picked out after G418 selection. Extraneous gene integration and expression were confirmed by PCR and semi-quantitative RT-PCR. The cytotoxicity to these transgenic cells under treatment with 5-Fc and GCV were measured by MT1 assays. Results Double and single suicide gene transfer were both stably expressed in GLC-82 cells . The cytotoxic effects of co-expressed TK-CD genes were superior than that of the single gene. Conclusion The CD + TK/5-Fc + GCV coexpression system is more effective for killing effect of tumor cells than CD/5-Fc or TK/GCV system alone.

AIMTo investigate the effect of brassinolide, a plant growth modulator, on multidrug resistance (MDR) of human T lymphoblastoid cell line CCRF-VCR 1000 which was obtained by progressively addition of vincristine (VCR) to sensitive CCRF-CEM cells, and to explore preliminarily the mechanism of reversing action.

METHODSMTT method was used to detect the resistant factor of resistant cell line and the reversing fold after addition of brassinolide. The intracellular accumulation of rhodamine 123, a fluorescent dye transported by P-glycoprotein was detected by flow cytometry, the catalytic activity of topoisomerase II was assessed by Sulliven method to find the effect of brassinolide on resistance. The protein expression of p53 was measured using Western blotting in the sensitive cells and resistant cells to explore the effect of brassinolide.

RESULTSThe resistant factors of CCRF-VCR cells on adriamycin, VP-16 and VCR are respectively as 153.1, 55.9 and 8123.1 folds comparing to the sensitive cell line CCRF-CEM. After treatment of brassinolide under the concentration of 0.001 - 10.0 microg x mL(-1), the resistance of CCRF-VCR was reversed partly with the reversing folds respectively as 4.4 - 11.6. The intracellular accumulation of rhodamine 123 was significantly reduced in the resistant cells. After treatment of brassinolide, the accumulation increased, the level of fluorescent dye was situated between resistant cells and sensitive cells. No alteration of the catalytic activity of topoisomerase II was found among three groups. The level of protein expression of p53 in resistant cells was higher than that of sensitive cells. After brassinolide treatment, the expression of p53 in CCRF-VCR cells restored to the level of sensitive cells.

CONCLUSIONBrassinolide could effectively reverse the resistance of CCRF-VCR cells by inhibiting the effusion of drug transported by P-glucoprotein. To down regulate the abnormal expression of p53 maybe one of the mechanisms of reversing MDR for brassinolide.

Brassica rapa ; chemistry ; Brassinosteroids ; Cell Line, Tumor ; drug effects ; Cholestanols ; isolation & purification ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Humans ; Leukemia, T-Cell ; metabolism ; pathology ; Plant Growth Regulators ; pharmacology ; Pollen ; chemistry ; Steroids, Heterocyclic ; isolation & purification ; pharmacology ; Tumor Suppressor Protein p53 ; metabolism

Orexin-A is a novel neuropeptide produced by neurons mainly located in lateral hypothalamic area that potently facilitates appetite and food intake. The purpose of this study was to investigate the possible change in orexin-A immunoreactivity in suckling-induced hyperphagia. By using immunohistochemistry and image analysis techniques we examined orexin-A-like immunoreactivity in a series of rat brain sections corresponding to the hypothalamus in groups of non-lactating, lactating, lactating with overnight cessation of suckling, lactating and cessation followed by resumed short-term sucklings. Long-term lactation significantly increased daily food intake on day 3 (81%) and day 11 (180%) postpartum compared to that in non-lactating postpartum rats, whereas daily food intake was significantly decreased by overnight cessation of suckling on day 11 postpartum in long-term lactating rats (45%). Moreover, long-term lactating rats on day 12 postpartum exhibited significantly greater number and higher mean staining intensity of orexin-A immunoreactive neurons than those of non-suckling postpartum rats (P<0.001 and P<0.05, respectively). Overnight cessation of lactation in rats on day 12 postpartum significantly decreased both the number and mean staining intensity of orexin-A immunoreactive neurons compared to those in long-term lactating group of rats (P<0.001 and P<0.05, respectively), similar to the levels in the non-lactating postpartum rats. Resumed lactation for 2 and 5 h after overnight cessation of lactation significantly increased the number (P<0.001 and P<0.05, respectively) and mean staining intensity (P<0.05) of orexin-A immunoreactive neurons compared to those in the rats without resumed lactation. Both long-term lactation and short-term resumed suckling enhanced orexin-A immunoreactivity in the hypothalamus in rats, and overnight cessation of lactation down-regulated the increased orexin-A immunoreactivity induced by long-term lactation. Suckling may regulate orexin-A expression in the hypothalamus and the increased orexin-A may be involved in hyperphagia in lactating rats, suggesting the possibility of the existence of some neural-humoral links between suckling and hypothalamic orexin-A-immunoreactive neurons.
Animals ; Feeding Behavior ; physiology ; Female ; Hyperphagia ; physiopathology ; Hypothalamus ; metabolism ; physiology ; Immunohistochemistry ; Intracellular Signaling Peptides and Proteins ; immunology ; metabolism ; Male ; Neurons ; metabolism ; physiology ; Neuropeptides ; immunology ; metabolism ; Neurotransmitter Agents ; immunology ; metabolism ; Orexins ; Rats ; Rats, Wistar

·AIM: To study the effect and safety of sodium hyaluronate combined with loteprednol eye drops compared to pure sodium hyaluronate eye drops in the clinical treatment of dry eye in children. ·METHODS: From August 2014 to August 2016, 128 children (256 eyes) with water deficient dry eye treated with artificial tears in our hospital were randomly divided into two groups, 64 cases (128 eyes) in each, the combined group and the control group, according to the random number table method. The combined group was given sodium hyaluronate combined with loteprednol treatment;the control group was given simple treatment of sodium hyaluronate eye drops. Before and after treatment for 2 and 6wk for ocular surface analysis system (Keratograph 5M) examination, determination of tear break-up time (BUT), tear meniscus height (TMH), meibomian gland infrared photography, slit lamp observation of corneal fluorescein staining, the observation indexes were statistically analyzed. ·RESULTS: After 2wk of treatment, the TMH and BUT level of the two groups were all higher than those before treatment(P<0.05). The scores of meibomian gland score and corneal fluorescein staining were lower than those before treatment, and the difference was statistically significant (P<0.05). After 6wk of treatment, the TMH and BUT level of the two groups were all higher than those before treatment and 2wk after treatment; The meibomian gland score and scores of corneal fluorescein staining were lower than those before treatment and 2wk after treatment, the difference was statistically significant (P<0.05). After 2wk and 6wk of treatment,TMH and BUT level combined groups were higher than those in the control group, while the score of meibomian gland and corneal fluorescein staining scores were lower than those in the control group, the difference was statistically significant (P<0. 05). Intraocular pressure between the two groups before and after treatment was not significantly different (P>0.05). · CONCLUSION: Sodium hyaluronate combined with loteprednol eye drops is more effective in the clinical treatment of dry eye children than the use of sodium hyaluronate eye drops,while the safety and tolerance are better.

Objective To evaluate the clinical effect of femtosecond laser assisted cataract surgery with Toric IOL implantation on correcting astigmatism and IOL rotating stability.Methods A prospective,nonrandomized,controlled study was conducted in 68 patients (82 eyes) diagnosed as age-related cataract with more than 1.0 diopter (D) regular astigmatism,including 41 eyes undergoing Toric IOL implantation with femtosecond laser assisted cataract surgery,and the other eyes receiving Toric IOL with phacoemulsification.And observation was performed for the uncorrected visual acuity (UCVA) before operation and 6 months after operation,preoperative corneal astigmatism,anticipated and postoperative residual astigmatism 6 months after operation.Then,comparison was performed for the degree of Toric lens axis rotation,as well as the horizontal and vertical movement of the IOL and the degree of vertical deflection in the capsule at 1,3,and 6 months after operation.Results In the observation group,preoperative UCVA (logMAR),preoperative corneal astigmatism,postoperative BCVA and the residual astigmatism at 6 months after operation was 1.03 ± 0.32,(2.09 ± 0.73) D,0.13 ± 0.11 and (0.33 ± 0.27) D,respectively,while the corresponding data in the control group was 0.92 ± 0.40,(1.69 ± 0.64) D,0.13 ± 0.09,(0.26 ± 0.22) D;the postoperative visual acuity in both groups was significantly improved,and residual astigmatism was significantly decreased at 6 months after operation (all P ＜ 0.001).There was no significant difference in Toric lens axis after femtosecond laser assisted surgery and phacoemulsification at 1 month,3 months,6 months after operation (all P ＞ 0.05),but the differences in the IOL movement in horizontal and vertical direction were statistically significant at the different time-points (all P ＜ 0.05),and there was significant differences in the degree of vertical deflection (all P ＜ 0.05).Conclusion Both femtosecond laser assisted cataract surgery and phacoemulsification combined with Toric IOL can achieve a better stability and predictability.

Objective To investigate the intervention effect of CD147 on learning and memory ability in rat model of Alzheimer's disease. Methods A total of 60 healthy Sprague-Dawley rats were randomly divided into sham operation group, model group and CD147 group, 20 rats in each group. All of the rats were anesthetized with intraperitoneal injection of 10％chloral hydrate (0.3 g/kg). The rats in the model group and the CD147 group were injected with Aβ1-40 (10μg) in the bilateral hippocampal CA1 regions, while the rats in the sham operation group were injected with the same amount of saline at the same sites. After 48 h, the rats in CD147 group were injected with CD147 cDNA in the bilateral ventricles, while the rats in model group and sham operation group were injected the same amount of saline at the same sites. Morris water maze test was used to detect the ability of learning and memory of rats. The expressions ofβamyloid protein (Aβ) andγ-secretase were detected by Western blot assay. Results The escape latency was significantly longer in model group than that of sham operation group, while which was significantly lower in CD147 group than that of model group (P<0.05). The number of times across the platform and the time of staying on platform were significantly lower in model group than those of sham operation group, while which was significantly higher in the CD147 group than that of model group (P<0.05). The expressions of Aβandγ-secretase were increased significantly in model group compared to those of sham operation group, while which were significantly decreased in CD147 group compared with those of model group (P<0.05). Conclusion Exogenous CD147 can significantly improve the learning and memory ability of AD rats, and its specific mechanism may be related to regulating the activity ofγ-secretase and down regulating the expression of Aβ.

OBJECTIVETo explore a simple, rapid and efficient way to generate dendritic cells from leukemic cells.

METHODSK562 cells were cultured with calcium ionosphere A23187 alone, A23187 plus GM-CSF, or a DC differentiation cocktail consisting of GM-CSF, IL-4 and TNF-alpha, respectively. The expression of surface markers of induced DCs was analyzed by flow cytometry. The K562-DCs stimulating the proliferation of allo-genetic naive T cells and inducing cytotoxicity of T cells were determined by MTT assay.

RESULTSMicroscopic examination revealed that under all the three culture conditions, K562 cells became displaying DC morphology. At 72 hours in the two culture systems containing A23187, there were higher proportions of cells with dendritic morphology [(69.5 +/- 17.2)% and (73.1 +/- 13.9)%, respectively] than that in the cocktail system [(28.5 +/- 12.3)%] (P < 0.05). And the same did when cultured for 7 days [(69.5 +/- 17.2)%, (73.1 +/- 13.9)% respectively vs (51.2 +/- 10.7)%, P < 0.05]. In the 7-day cultures, the percentage of CD1a expressing cells was lower [(8.2 +/- 2.3)% and (10.3 +/- 5.1)% vs (17.2 +/- 1.6)%, respectively] while the CD83 expressing cells was higher [(85.6 +/- 8.8)% and (82.4 +/- 9.1)% vs (77.4 +/- 12.9)%, respectively] compared with that in the cocktail system (P < 0.05). No significant difference was found in the allogeneic T cell proliferation response and induced T cell cytotoxicity between A23187 containing and cocktail groups (P > 0.05).

CONCLUSIONSA23187 treatment is a simple, rapid and efficient in vitro strategy for inducing dendritic cell from leukemic cells.

Calcimycin ; pharmacology ; Cell Differentiation ; drug effects ; Cells, Cultured ; Coculture Techniques ; Dendritic Cells ; cytology ; immunology ; metabolism ; Humans ; K562 Cells ; cytology ; drug effects

OBJECTIVETo investigate the effect of particulate cancellous bone impaction grafting in combination with total hip arthroplasty (THA) for acetabular reconstruction in patients with posttraumatic arthritis and bone loss after acetabular fractures.

METHODSTotally 15 consecutive cases with unilateral acetabular fracture were treated with bone impaction grafting in combination with THA in our department. There were 10 males and 5 females with mean age of 48.2 years (ranging from 36 to 73 years). Eight cases had the fracture at left hips, 7 at right hips. The average age at injury was 28 years (ranging from 18 to 68 years). The mean follow-up period was 4.3 years (ranging from 2 to 7 years).

RESULTSCompared with mean 42 points (ranging from 10 to 62) of the preoperative Harris score, the survival cases at the final follow-up had mean 84 points (ranging from 58 to 98). One patient had mild pain in the hip. No revision of the acetabular or femoral component was undertaken during the follow-up. Normal rotational centre of most hips was recovered except 2 cases in which it was 0.8 mm higher than that in opposite side. All of them had a stable radiographic appearance. Progressive radiolucent lines were observed in I, III zones in 2 cases. One patient had a nonprogressive radiolucent line in zone III. The cup prosthesis was obviously displaced (6 mm) in one patient, but had not been revised.

CONCLUSIONParticulate cancellous bone impaction grafting in combination with THA as a biological solution is an attractive procedure for acetabular reconstruction in patients with posttraumatic arthritis and bone loss after acetabular fracture, which can not only restore acetabular bone stock but also repair normal hip anatomy and its function.

Acetabulum ; surgery ; Adolescent ; Adult ; Aged ; Arthritis ; surgery ; Arthroplasty, Replacement, Hip ; Bone Substitutes ; Bone Transplantation ; methods ; Female ; Hip Fractures ; surgery ; Humans ; Male ; Middle Aged

OBJECTIVETo evaluate the correlation between thrombosis and stability of atherosclerotic plaque within criminal vessels in patients with unstable angina pectoris (UAP) by coronary angioscopy, to explore the clinical pathological basis for acute coronary syndromes (ACS).

METHODSSixty-eight patients with UAP were enrolled, the patients with post-infarction angina pectoris and variant angina pectoris were excluded. There were 48 males and 20 females, aged from 40 to 73 (average 62.4 +/- 8.6) years. The criminal vessels of there patients were observed by coronary angioscopy during percutaneous coronary intervention (PCI) therapy.

RESULTSThere were 68 criminal vessels in 68 patients. Atherosclerotic plaques were observed in all criminal vessels. Among criminal vessels, thrombi and intimae lesions were detected in 63 cases and 46 cases, respectively. Among 68 cases with atherosclerotic plaques, there were 48 cases of yellow plaques (70.5%), 18 cases of light yellow plaques (26.5%) and 2 cases of white plaques (2.94%). Sixty-three thrombi cases were mural and on-occlusive, which included 11 cases of red or mixed thrombi (17.5%) and 52 cases of white or pink thrombi (82.5%). All intimae lesions were accompanied by thrombosis, which included 11 cases of red or mixed thrombi (23.9%) and 35 cases of white or pink thrombi (76.1%).

CONCLUSIONThe study has shown that the rupture of unstable yellow plaque and its thrombosis were the pathological basis of UAP. Therefore, stabilizing yellow plaque before its rupture may play critical role in prevention and treatment of ACS.

Adult ; Aged ; Angina, Unstable ; pathology ; Angioscopy ; Coronary Artery Disease ; etiology ; pathology ; Coronary Thrombosis ; etiology ; pathology ; Female ; Humans ; Male ; Middle Aged

The recombined adenovirus DNA was transfected into 293 cells for packing and amplification of replication-deficient Ad-CMV-E6/E7, Ad-K14 -E6/E7 virus was purified by CsCl density gradient centrifugation , recombined adenovirus Ad-CMV-E6/E7, Ad-K14 -E6/E7 were used as experimental group, while pAd-CMV and pAdtrack-K14 were used as control group. Four of them were injected through one main vein of nude mice tail respectively. These mice were then treated with 0.05 mg 17beta-estradiol over 12 weeks. Mice were anaesthesiaed with 2.5% Avertint and the vagina, mammary gland, ovaries and uterus were dissected and fixed in 3.75% paraformaldehyde overnight at 4 degrees C. Paraffin-embedded sections, HE staining and identification of P53 and Bcl-2 protein via immunohistochemistry were performed. The expression of E6/E7 was verified by RT-PCR in different tissue of nude mice. HE staining showed evident hyperplasy in cervix-uterus transformation zone of experimental group 2. The expression of mutant P53 and Bcl-2 were higher than control group via immunohistochemical S-P method in uterus stroma-cell. Western blotting also showed that E6 protein was expressed. The expression of E6/E7 was higher than control group by human cytokeratin promoter 14 and hyperlasy changes were detected in epithelial tissue of cervix-uterus transformation zone.
Adenoviridae ; genetics ; Animals ; Blotting, Western ; Cell Line ; Female ; Genital Diseases, Female ; pathology ; virology ; Genitalia, Female ; pathology ; virology ; Humans ; Immunohistochemistry ; Mammary Glands, Animal ; metabolism ; pathology ; Mice ; Mice, Nude ; Oncogene Proteins, Viral ; genetics ; metabolism ; Ovary ; metabolism ; pathology ; Papillomaviridae ; metabolism ; physiology ; Papillomavirus E7 Proteins ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Repressor Proteins ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Tumor Suppressor Protein p53 ; metabolism ; Uterus ; metabolism ; pathology ; Vagina ; metabolism ; pathology