Objective To observe the effect of rhTRAIL on survivin gene expression of human lung adeno-carcinoma A549 xenografted tumor in nude mice,and investigate the possible inhibitory mechanism of rhTRAIL on the implanted-tumor growth.Methods The solid tumor model was formed in nude mice with human lung adeno-carcinoma cell line.A549.24 mice were randomly divided into the four groups,rhTRAIL single treated group (1 μg/mL),rhTRAIL combined with cisplatin (DDP) treated group,cisplatin treated (1.5mg/kg) and 0.9％ sodium chloride injection(NS) control group.The rhTRAIL and DDP were injected once every other day by intraperitoneal injection to mice in the treated groups,lasting eight times,the same volume of saline solution was injected to the control group.After these,mice were killed and dissected completely.The expression level of survivin mRNA and protein in the tumor tissues was detected by real-time polymerase chain reaction (RT-PCR) and immunohistochemistry,respectively.And the expression of survivin gene in serum of each group was tested by ELISA.Results The expression levels of survivin mRNA in implanted-tumor tissues in rhTRAIL,rhTRAIL combined with DDP,DDP and NS group were (48.7 ± 2.5) ％,(53.1 ± 4.6) ％,(99.1 ± 5.3) ％ and (95.6 ± 3.7) ％,respectively.While the protein expressions of survivin gene in those groups were (0.319 ± 0.025),(0.483 ± 0.058),(0.635 ± 0.041) and (0.619 ± 0.017),respectively.Moreover,the serum levels of survivin were (71.9 ± 7.05),(80.26 ± 10.80),(112.75 ± 15.41) and (105.03 ± 20.37),respectively.The data showed that the expression levels of rhTRAIL and rhTRAIL combined with DDP group were lower than that of DDP-treated group or the NS control group (P ＜ 0.0 5).Compared with the rhTRAIL combined with DDP group,the survivin gene expression level of rhTRAIL-single treated group decreased a little lower,but the difference was not significant (P ＞ 0.05).Conversely,the survivin gene level was increased to some degree compared with the NS control group,and uniformly there was no significant difference (P ＞ 0.05).Conclusion rhTRAIL can downregulate the expression level of survivin gene of human lung adeno-carcinoma A549 xenografted tumor in nude mice.It may be one of the possible inhibitory mechanisms of rhTRAIL on the implanted-tumor growth that rhTRAIL can downregulate survivin gene expression and promote tumor cell apoptosis.

OBJECTIVE:To determine the content of IL-2 in IL-2 targeting liposomes of 5-FURP. METHODS: The sample solution was stained by Coomassie brillant blue G-250 solution. Using blank liposome as blank,the absorbability of the IL-2 was determined by ultraviolet spectrophotometry at a wavelength of 595 nm;meanwhile its content was determined. RESULTS: The linear range of IL-2 was 1～100 ?g?ml-1(r=0.999 7) and the average recovery rate was 99.26% (RSD=0.59%,n=5). CONCLUSION: The method was proved to be simple,rapid,sensitive and reproducible,and it is applicable for the determination of IL-2 in liposome of 5-FURP.

BACKGROUND:The researches about the effect of retinoic acid on the proliferation of adipose-derived stem cells are rare, and the researches on the testosterone are mainly on the inhibition of cellaging. OBJECTIVE: To study the effects of retinoic acid and testosterone or combination on the cellcycle of adipose derived stem cells. METHODS:Adipose derived stem cells were isolated from adult female Sprague Dawley rats with 2 months age and cultured in vitro til passage 3 adipose derived stem cells, and then the 3rd passage adipose-derived stem cells were performed with adipogenic induction, osteogenic induction and surface marker identification. The cells were divided into six groups:(1) Control group;(2) 10-5 mol/L retinoic acid group;(3) Retinoic acid group;(4) 10-5 mol/L retinoic acid+testosterone group;(5) 10-6 mol/L retinoic acid+testosterone group;(6) Testosterone group. The adipose-derived stem cells in the control group were cultured with Dulbecco’s modified Eagle’s medium+10%fetal bovine serum culture medium, and the adipose-derived stem cells in the other five groups were induced with corresponding dose of retinoic acid and testosterone on the basis of control group. After cultured for 36 hours, the flow cytometry was used to detect the changes of cellcycle. RESULTS AND CONCLUSION:Compared with the control group, cellproportions in phase G 1 of 10-5 mol/L retinoic acid group and 10-6 mol/L retinoic acid group were increased significantly, and the cellproportions in phase S were decreased. Compared with control group, the cellproportion in phase G 1 of testosterone group was significantly reduced, and the cellproportion in phase S was increased. Compared with 10-5 mol/L retinoic acid group and 10-6 mol/L retinoic acid group, cellproportions in phase G 1 of 10-5 mol/L retinoic acid+testosterone group and 10-6 mol/L retinoic acid+testosterone group were reduced significantly and the cellproportions in phase S were increased. Retinoic acid can inhibit the cellcycle of adipose-derived stem cells in phase G 1 , and delay the process of the cellcycle from phase G1 to phase S;while testosterone can promote the cellcycle of adipose-derived stem cells from phase G1 to phase S;the combination induction of retinoic acid and testosterone can accelerate the process of the cellcycle of adipose-derived stem cells from phase G 1 to phase S.

Objective To evaluate the level changes of serum WASP-family verprolin homologous protein-1 (WAVE1) and vascular endothelial growth factor-C (VEGF-C) and their clinical significance in patients with advanced non-small lung cancer (NSCLC) before and after chemotherapy.Methods Serum WAVE1 and VEGF-C were measured in 43 patients with advanced NSCLC by ELISA,and the results were compared with 43 healthy volunteers.Results The levels of serum WAVE1 and VEGF-C before chemotherapy in patients group were (0.573±0.082) ng/ml and (947.3±125.4) pg/ml respectively,while in healthy volunteers group,they were (0.256±0.064) ng/ml and (425.5±110.1) pg/ml respectively,which suggested that before chemotherapy the levels of serum WAVE1 and VEGF-C in NSCLC group were significantly higher than those of in the control (P ＜ 0.05).The serum levels of WAVE1 and VEGF-C in advanced NSCLC patients were closely related to lymph node metastasis status and distant metastasis status (P ＜ 0.05),but not to the gender,age,tumor length,histology type,differentiation grade and C-TNM stage (P ＞ 0.05).The serum WAVE1 and VEGF-C levels of the effective treatment group was (0.290±0.037) ng/ml and (596.1±127.5) pg/ml after chemotherapy respectively,which decreased obviously compared with the group before chemotherapy which levels were (0.517±0.051) ng/ml and (964.6±100.3) pg/ml (both P ＜ 0.05).But the serum WAVE1 and VEGF-C levels of the ineffective treatment group were (0.547±0.065) ng/ml and (957.0±111.2) pg/ml after treatment,which had no difference compared with the group before chemotherapy which levels were (0.517±0.051) ng/ml and (964.6±100.3) pg/ml (both P ＞ 0.05).Furthermore,statistically significant relationship was found between the serum WAVE1 and the VEGF-C levels (r =0.331,r =0.540,both P ＜ 0.05).Conclusion Serum WAVE1 and VEGF-C may be used as indicators for prediction of the efficacy of chemotherapy in patients with advanced NSCLC.

Objective To study the therapeutic effect and safety of pleurodesis with medical thoracoscopy by powdery talc on treatment of malignant pleural effusion.Methods Retrospective analysis were done in 74 cases of malignant pleural effusion accepted simple powdery talc pleurodesis under medical thoracoscopy.Results The doses of powdery talc were from 1g to 5g,average 2.18g in the course of the treatment.After operation,45 (60.8％) cases which were complete remission(CR) were full pleural adhesion and complete lung recruitment,14cases (18.9％),which were partial remission(PR),were less pleural adhesion and most lung recruitment,and 10cases(13.6％) were stable diseases(SD),while 5cases(6.7％),which were progressive diseases(PD),were without pleural reaction.The total effective rate,including CR,PR and SD,was 93.3 ％ (69/74).The complications of simple powdery talcage under medical thoracoscopy were included in:95.9％ (71/74) with chest pain,64.8％ (48/74) with fever,28.4％ (21/74) with shortness of breath,12.2 ％ (9/74) with mediastinal and subcutaneous emphysema,5.4％ (4/74) with nausea and vomiting,4.1％ (3/74) with abdominal distension,while the complications of acute pulmonary edema,massive hemorrhage,pulmonary embolism and wound infection were not observed.Conclusion Treatment of malignant pleural effusion by simple powdery talcage under medical thoracoscopy has definite clinic therapeutic effect,which is safe,cost-effective,less trauma and worthy of clinical application.

Objective This study aimed to investigate the prevalence of Epstein-Barr virus (EBV) infection in patients with and without cancer. Methods A total of 26,648 participants who underwent whole-blood EBV DNA (WBEBV) assays between January 1,2020,and August 31,2023,were included. The chi-square test was used for categorical data analysis,and R software was used to analyze the differences in EBV DNA load levels and the diagnostic capabilities of WBEBV. Results Positive rates were 10.2％ and 25.4％ for healthy controls (HC) and patients,respectively. The positivity rate for EBV-associated neoplasms (EN) was the highest at 7.53％,followed by leukemia (Le) at 5.49％. The subgroup analysis showed that the positivity rate for abnormal proliferation or hyperplasia (APH) was 31.9％,followed by 30.5％ for Le. The WBEBV of patients with transplants (TP),especially living-related transplants (LT),was the highest among all subgroups. WBEBV at diagnosis was used to differentiate between infectious mononucleosis (IM) and chronic active Epstein-Barr virus (CAEBV),with a sensitivity of 67.4％ (95％ confidence interval[CI]:57.6-75.8) and specificity of 72％ (95％ CI:63.3-79.3). We conclude that the prevalence of EBV infection is low in the healthy population in this region and that a high EBV load at baseline is more common in LT,IM,and Lymphocyte Leukemia (LL). Conclusion This study used a large-sample survey to characterize the prevalence of whole-blood EBV levels in various diseases,including the stages and subtypes. The EBV detection rate was higher in patients with malignant disease than in those with benign disease. Our study provides clinicians with baseline information regarding EBV-associated diseases.

OBJECTIVETo investigate whether adipose-derived stem cells (ADSCs) induced by retinoic acid (RA) in vitro express primordial germ cell marker alkaline phosphatase (ALP) and vasa.

METHODSADSCs were isolated from adult female SD rats and cultured in vitro. The third passage of ADSCs was identified by adipogenic differentiation, osteogenic differentiation and cell surface marker detection. The ADSCs were treated with 1×10(-5), 1×10(-6), or 1×10(-7) mol/L RA for 7 or 14 days, and the cellular expression of ALP was detected. vasa mRNA expression in ADSCs treated with 1×10(-5) mol/L RA for 7 days was detected using RT-PCR.

RESULTSThe OD value of ADSCs treated with 1×10(-5), 1×10(-6), or 1×10(-7) mol/L RA was 0.59∓0.04, 0.27∓0.07, and 0.15∓0.03 after a 7-day treatment, and was 0.42∓0.02, 0.34∓0.01, and 0.19∓0.02 after a 14-day treatment, respectively, demonstrating significantly enhanced ALP expression in RA-treated ADSCs compared with that in the control cells (0.07∓0.01 and 0.07∓0.01 at 7 and 14 days, respectively, P<0.01). The ADSCs showed a negative vasa mRNA expression after 1×10(-5) mol/L RA treatment for 7 days.

CONCLUSIONRA-induced ADSCs express ALP, a marker of primordial germ cells, but does not express the primordial germ cell marker vasa.

Adipose Tissue ; cytology ; Adult Stem Cells ; cytology ; enzymology ; Alkaline Phosphatase ; metabolism ; Animals ; Cell Differentiation ; Cells, Cultured ; Female ; Germ Cells ; cytology ; metabolism ; Rats ; Rats, Sprague-Dawley ; Tretinoin ; pharmacology

Jingzhaotoxin-V(JZTX-V) isolated from the venom of the spider Chilobrachys jingzhao is a novel potent inhibitor that acts on tetrodotoxin-resistant and tetrodotoxin-sensitive sodium channels in adult rat dorsal root ganglion(DRG) neurons. It is a 29-residue polypeptide toxin including three disulfide bridges. To investigate the structure-function relationship of the toxin, a mutant of JZTX-V in which Arg20 was substituted by Ala, was synthesized by solid-phase chemistry method with Fmoc-protected amino acids on the PS3 automated peptide synthesizer. The synthetic linear peptide was then purified by reversed-phase high performance liquid chromatography and oxidatively refolded under the optimal conditions. The refolded product was analyzed by matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry(MALDI-TOF MS) and electrophysiological experiments for its relative molecular weight and prohibitive activity of sodium channels respectively. The present findings show that the prohibitive effect of R20A-JZTX-V on TTX-S sodium channels in DRG neurons is almost the same as that of native JZTX-V, suggesting that Arg20 does not play any important role in inhibiting TTX-S sodium currents in DRG neurons. In contrast, the prohibitive level of R20A-JZTX-V on TTX-R sodium channels is reduced by at last 18.3 times, indicating that Arg20 is a key amino acid residue relative to the bioactivity of JZTX-V. It is presumed that the decrease in activity of R20A-JZTX-V is due to the changes of the property in the binding site in TTX-R sodium channels.
Amino Acid Substitution ; Animals ; Arginine ; genetics ; Ganglia, Spinal ; drug effects ; Mutagenesis, Site-Directed ; Mutant Proteins ; pharmacology ; Neurons ; drug effects ; Patch-Clamp Techniques ; Peptides ; chemistry ; genetics ; pharmacology ; Rats ; Sodium Channel Blockers ; pharmacology ; Sodium Channels ; drug effects ; Spider Venoms ; chemistry ; genetics ; isolation & purification ; pharmacology ; Spiders ; Tetrodotoxin ; pharmacology

Kv2.1 channel currents in pancreatic beta-cells are thought to contribute to action potential repolarization and thereby modulate insulin secretion. Because of its central role in this important physiological process, Kv2.1 channel is a promising target for the treatment of type 2 diabetes. Jingzhaotoxin-XI (JZTX-XI) is a novel peptide neurotoxin isolated from the venom of the spider Chilobrachys jingzhao. Two-microelectrode voltage clamp experiments had showed that the toxin inhibited Kv2.1 potassium currents expressed in Xenopus Laevis oocytes. In order to investigate the structure-function relationship of JZTX-XI, the natural toxin and a mutant of JZTX-XI in which Arg3 was replaced by Ala, were synthesized by solid-phase chemistry method with Fmoc-protected amino acids on the PS3 automated peptide synthesizer. Reverse-phase high performance liquid chromatography (RP-HPLC) and matrix assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF/TOF MS) were used to monitor the oxidative refolding process of synthetic linear peptides to find the optimal renaturation conditions of these toxins. The experiments also proved that the relative molecular masses of refolded peptides were in accordance with their theoretical molecular masses. RP-HPLC chromatogram of co-injected native and refolded JZTX-XI was a single peak. Under the whole-cell patch-clamp mode, JZTX-XI could completely inhibit hKv2.1 and hNav1.5 channels currents expressed in HEK293T cells with IC50 values of 95.8 nmol/L and 437.1 nmol/L respectively. The mutant R3A-JZTX-XI could also inhibit hKv2.1 and hNav1.5 channel currents expressed in HEK293T cells with IC50 values of 1.22 micromol/L and 1.96 micromol/L respectively. However, the prohibitive levels of R3A-JZTX-XI on hKv2.1 and hNav1.5 channels were reduced by about 12.7 times and 4.5 times respectively, indicating that Arg3 was a key amino acid residue relative to the hKv2.1 channel activity of JZTX-XI, but it is also an amino acid residue correlated with the binding activity of JZTX-XI to hNav1.5 channel. Our findings should be helpful to develop JZTX-XI into a molecular probe and drug candidate targeting to Kv2.1 potassium channel in the pancreas.
Animals ; HEK293 Cells ; Humans ; Insulin-Secreting Cells ; metabolism ; Mutant Proteins ; genetics ; pharmacology ; NAV1.5 Voltage-Gated Sodium Channel ; metabolism ; Neurotoxins ; chemical synthesis ; genetics ; pharmacology ; Protein Refolding ; Shab Potassium Channels ; antagonists & inhibitors ; metabolism ; Sodium Channel Blockers ; pharmacology ; Spider Venoms ; genetics ; pharmacology ; Transfection