Objective To investigate the effect of cold stimulation on the phenotype of alveolar macrophages(MH-S cells) in mice. Methods MH-S cells were cultured at 37 ℃ for 24 h,and cold stimulated at 36,34 and 32 ℃ for 0,0. 5,1,3,6,9 and 12 h respectively. The mRNA transcription levels of interleukin-1β(IL-1β) and interleukin-10(IL-10) genes in MH-S cells were detected by qRT-PCR. MH-S cells were cultured at 37 ℃ for 24 h,and cold stimulated at 34 ℃ for 0. 5 h,which were detected for the mRNA transcription levels of tumor necrosis factor-α(TNF-α),inducible nitric oxide synthase(iNOS)and Arginase1(Arg1)genes by qRT-PCR(MH-S cells with 0 h cold stimulation as control),detected for the expression of iNOS and Arg1 by immunofluorescence assay(MH-S cells cultured at 37 ℃ for 0. 5 h as negative control)and detected for the expression levels of iNOS,TNF-α and nuclear factor-kappa B(NF-κB)by Western blot(MH-S cells cultured at 37 ℃ for 0. 5 h as negative control). Results The mRNA transcription levels of IL-1β and IL-10 genes in MH-S cells were the highest when the cells were cultured at 34 ℃ for 0. 5 h,therefore,the cold stimulation model of MH-S cells was established under this condition. Compared with the cells cultured for 0 h,the mRNA transcription levels of iNOS,TNF-α and Arg1genes in MH-S cells cultured at 34 ℃ for 0. 5 h increased significantly(t = 3. 733,12. 190 and 6. 793,respectively,each P < 0. 05). Compared with the negative control group,the fluorescence expression intensity of iNOS and Arg1 in MH-S cells in the stimulation group increased,especially iNOS,the expression levels of iNOS and TNF-α proteins increased with no significant difference(t = 0. 675 and 1. 514,respectively,each P > 0. 05),and the expression level of NF-κB increased significantly(t = 3. 092,P < 0. 05). Conclusion Cold stimulation at 34 ℃ for 0. 5 h can increase the expression of inflammatory factors such as IL-1β,IL-10,TNF-α,iNOS,Agr1 and NF-κB in MH-S cells,activate NF-κB signaling pathway in MH-S cells,induce the expression of inflammatory proteins and promote cell activation.

OBJECTIVE:To study the separation and purification of polysaccharides sulfate from Ulva lactuca,and to analyze its structure.METHODS:The separation and purification method of polysaccharides sulfate from U.lactuca was optimized.The content of polysaccharides in polysaccharides sulfate was determined by phenyl-sulfuric acid method.The molecular amount of polysaccharides was determined by HPGC.The structure of polysaccharides was analyzed by IR and 13C-NMR.RESULTS:The optimal extraction condition of polysaccharides sulfate from U.lactuca were crashed materials 20 mesh sieve,20-fold water,decocting 3 times at 85 ℃,2 hours each time.The content of polysaccharides was 22.64% and molecular amount was 1 156 112.Soluble polysaccharides mainly contained ?-glucuronic acid,rhamnose and plenty of sulfate radical.CONCLUSION:The method is simple and stable,which provides theoretic basis for production of polysaccharides sulfate from U.lactuca.

BACKGROUND:Mesenchymal stem cells are found to have the immunoregulatory activities and a potential application prospect in the treatment of autoimmune diseases.
OBJECTIVE:To explore the mechanism of transplanting mesenchymal stems cells on the treatment of multiple sclerosis.
METHODS:The mouse mesenchymal stems cells were prepared, and injected into the al ogenic and syngenic normal mice, to detect the frequency of CD4+CD25+Foxp3+T cells in the spleen, thymus, and lymph nodes by flow cytometry, and to detect the Foxp3, transforming growth factor-β1, and interleukin-10 mRNA in the spleen, thymus, and lymph nodes by reverse transcription-PCR.
RESULTS AND CONCLUSION:Transplantation of mesenchymal stem cells on normal mice led to a significant up-regulation of CD4+CD25+Foxp3+T cells, Foxp3, transforming growth factor-β1, and interleukin-10 mRNA in the spleen, thymus, and lymph nodes both in the al ogenic and syngenic transplant groups. Transplantation of mesenchymal stem cells may be an available method in the treatment of autoimmune diseases, and CD4+CD25+Foxp3+T cell, Foxp3, transforming growth factor-β1, and interleukin-10 may be involved in this process.

Objective To compare the effect of percutaneous intraluminal radiofrequency ablation ( RFA ) combined with biliary stenting and that of percutaneous transhepatic puncture combined with biliary stenting .Methods A total of 56 patients with unresectable malignant obstructive jaundice were reviewed retrospectively .Among them, 25 patients had received percutaneous intraluminal RFA combined with biliary stenting ( RFA group) while another 31 patients had been simultaneously selected for the simple biliary stent implantation ( stent group ) .The changes of the serum total bilirubin ( TB) and direct bilirubin ( DB) before and after 7-14 days of treatment , surgical complications , stent median patency and the median survival were observed.Follow-up information was obtained through telephone reviews or check-up records. Results The technical success rate was 100%.No procedure-related peritonitis or perforation occurred .There were respectively 3 cases with cholangeitis in RFA group and 3 in stent group.All the cases was controlled by effective clinical treatment.There was obvious statistically significant difference after treatment in TB and DB in the two groups (P<0.01, P<0.01).TB and DB fell by (149.05 ±110.71) and (96.93 ±69.12)μmol/L after 7-14 days in RFA group vs (151.40 ±94.47) and (94.21 ±67.36)μmol/L in stent group.The changes of the two groups were of no statistical significances .The stent patenmedian time was 122 ( 9 -550 ) and 157 ( 16 -510 ) d, while the median survival was 125(9-550) and 163 ( 16 -520 ) d.The difference was of no statistical significance .Conclusion Percutaneous intraluminal RFA combined with biliary stenting and percutaneous transhepatic puncture combined with biliary stenting are both safe and feasible therapeutic options for unresectable malignant obstructive jaundice .There is no statistically significant diffference between the two groups in the recent and long-term curative effects .

Objective:To investigate the serum hepcidin level and risk factors associated with peripheral arterial disease (PAD) and foot ulcer in type 2 diabetic patients.Methods:From January 2019 to June 2019, 70 patients with type 2 diabetes in Department of Endocrinology of Xiangya Third Hospital were selected, including 21 newly diagnosed patients with type 2 diabetes (DM group), 23 patients with lower extremity vascular disease (PAD group) and 26 patients with foot ulcer (DF group). Serum hepcidin was determined by enzyme linked immunosorbent assay (ELISA). The serum levels of hepcidin in different groups were compared, and the correlation between diabetic lower extremity vascular disease and foot ulcer was analyzed.Results:⑴ The hemoglobin, albumin, triglycerides and total cholesterol were significantly lower in DF group compared with PAD and DM groups ( P<0.05), while the DF group patients were with higher white blood cell (WBC) count and high sensitivity C reactive protein (hs-CRP) than patients in PAD and DM groups ( P<0.05). DF group also showed significantly higher WBC, hs-CRP and neutrophil ratio level (NEUT%) than DM group ( P<0.05). The inflammatory indicators of WBC, hs-CRP and NEUT% showed no significant difference between DM group and PAD group ( P>0.05). ⑵ The levels of hepcidin in DF and PAD groups were higher than those in DM group, while that in DF group were higher than those in PAD group ( P<0.05); Hepcidin was positively correlated with systolic blood pressure, WBC count, NEUT% and ferritin ( P<0.05), and negatively correlated with hemoglobin, glycosylated hemoglobin, albumin and 25-hydroxyvitamin D ( P<0.05). ⑶ Binary multivariate logistic regression analysis showed that elevated hepcidin level was an independent risk factor for diabetic foot ulcer [ OR=1.755, 95% CI: 1.063-2.897, P=0.028]. Conclusions:The fluctuation of serum hepcidin level in diabetic patients is related to the stimulation of inflammation, the degree of anemia and the nutritional status, which means it might be an early indicator of inflammation in diabetic patients with peripheral arterial disease. Moreover, the increase of hepcidin is an independent risk factor for diabetic foot ulcers in our study.

Objective:To develop a biomimetic nanoparticle probe of hematoporphyrin monomethyl ether (HMME) coated with breast cancer cell membrane, to observe its ability to target homologous breast cancer cells in vitro, and to investigate its effect of enhanced photoacoustic imaging and sonodynamic therapy (SDT) for breast cancer in vitro.Methods:The cell membrane of breast cancer 4T1 was extracted by chemical cleavage and repeated freezing and thawing. Then the HMME-coated polylactic acid-glycolic acid copolymer biomimetic nanoparticle was prepared by double emulsification and extrusion. The basic characteristics of nanoparticles were detected. The target ability of nanoparticles to homologous breast cancer cells and the enhancement of photoacoustic imaging were observed in vitro. Singlet oxygen sensor green (SOSG) was used to verify the reactive oxygen species (ROS) production of nanoparticles, and its SDT effect on breast cancer cells was evaluated by CCK8 cytotoxicity assay.Results:The size of the prepared CHP-NPs was uniform, the morphology was spherical "core-shell structure" , the particle size was (275.23±8.25)nm, and the surface potential was (-18.43±0.45)mV. It was observed that CHP-NPs could target homologous 4T1 cells under laser confocal microscopy. In vitro photoacoustic imaging experiments show that the photoacoustic signal of nanoparticles increases with the increase of its concentration. According to SOSG probe detection, CHP-NPs could produce ROS under ultrasonic irradiation.When CHP-NPs was incubated with 4T1 cells alone and no ultrasonic irradiation was used, the cell survival rate was not significantly affected. When the concentration was 0.6 mg/ml, the cell survival rate was still 95%. After ultrasonic irradiation, CCK8 experiment showed that the CHP-NPs had a significant SDT effect on breast cancer cells.Conclusions:The biomimetic nanomolecular probe of breast cancer cell membrane is successfully prepared. The probe has good ability to target homologous tumor, and can significantly enhance tumor photoacoustic imaging and SDT effect.

BACKGROUND: T cells are believed to play an important role in anti-infection, anti-tumor and immune function. However, the mechanism underlying the differentiation and development remains poorly understood. OBJECTIVE: To investigate the distribution of T cells in nude mice that are jointly transplanted human thymus and cord blood and the reconstruction of the immune function. METHODS: Thirty Balb/c nu/nu nude mice were randomly divided into two groups: an experimental group and a control group. In the experimental group, human thymus tissue was transplanted into the renal capsule of nude mice. Two weeks later, freshly isolated human cord blood CD34+ cells suspension was back perfused into the nude mice via the vein. In the control group, CD34+ cells transplantation was performed directly without thymus transplantation. After 60 days of breeding, the immune function of nude mice was detected in two groups. RESULTS AND CONCLUSION: Human thymus tissue in the renal capsule of nude mice survived and expressed CD3 and HLA-DR molecule. In the experimental group, CD3+ cells which distributed in the form of dots were observed in the mouse spleen. The proportion of CD3+, CD4+, CD8+, and CD4+CD25+ cells were significantly higher in the experimental group than in the control group. The nude mice from the experimental group rejected human gastric cancer BGC823 cells, while those from the control group did not. These findings demonstrated that combined human thymus and CD34+ cell transplantation allow nude mice to acquire T cell-mediated cellular immune function and possess the ability of anti-tumor.

BACKGROUND: Further studies are needed to understand the cytobiological character, functional regulation, gene changes and expression difference of CD34~+ and CD133~+ stem cells induced by basic fibroblast growth factor (bFGF) using gene chip. OBJECTIVE: To compare the differences of gene expression and the response to bFGF of human umbilical cord CD34~+ and CD133~+ cells, and to explore gene expression changes of bFGF-induced umbilical cord CD34~+ and CD133~+ hematopotic stem cells/hemapoietic progenitor cells in vitro. METHODS: Human umbilical cord blood CD34~+ and CD133~+ cells were isolated and purified by MiniMACS immunomagnetic beads selection. The CD34~+ and CD133~+cells were cultured for 10 to 15 days in DMEM/F12 medium, supplemented with bFGF and B27. Total RNA from these cells was extracted and the genetic level of these cells was performed using Oligo GEArray(r) chip and GEArray software. Selected rate of CD34~+ and CD133~+ hematopoietic stem cells was detected using flow cytometry. CD34~+ and CD133~+ cell morphological changes were measured before and after bFGF induction. The concentration and purity of RNA were determined by agarose gel electrophoresis degeneration. Gene-chip test results were analyzed.

Objective To study the over-expression and clinical implications of the oncogene MDM2 in acute leukemia (AL). Methods The expression of MDM2 gene in 100 patients with newly diagnosed and relapse or refractory AL and 20 healthy as control was measured by relative quantitative reverse transcriptase polymerase chain reaction (RT-PCR),then the results was measured by χ2-test,t-test and one-way ANOVA to compare expession positive rate and intensity of MDM2. Results Among 100 patients,fifty-eight had the high expression of MDM2 gene (58 %). The expression level of MDM2 gene in patients was higher than that of health controls(P ＜0.05). The expression positive rate of MDM2 is higher in poor outcome group (67.9 %,19/28)than that in general outcome group (33.9%,19/56) (P＜0.05). Conclusion Our results suggest that the expression of MDM2 gene plays an important role in the pathogenesis and poor outcome of AL.

Objective To investigate ATM deletion [del (ATM)] in chronic lymphocytic leukemia (CLL) and study its correlation with the clinical stage. Methods Spectrum Orange~(TM) labeled sequence specific DNA probe for ATM locus on 11q22.3 and fluorescence in Situ hybridization (FISH) was used to examine del (ATM) in 28 newly diagnose patients with CLL. FISH analysis were performed on bone marrow smears. Clinical staging was done according to Binet Method.Fisher exact propability was used to study the relations between del (ATM) and clinical feature. Results 4 out of the 28 cases were found with deletion of ATM. The incidence of del (ATM) in BinetA, BinetB and BinetC was 1/9 (11.1 %), 1/8 (12.5 %), 2/11 (18.2 %), respectively. Fisher exact propability showed that deletion of ATM was not associated with its clinical feature. Conclusion Application of FISH on archival bone marrow smears is a simple, liable method, and can be readly used to retrospective study of clonal blood system diseases. Deletion of ATM was common cytogenetic changes in CLL patients.And the significance of del (ATM) in the prognosis of CLL in China needs to be further investigated.