AIM To prepare diosgenin nanosuspensions.METHODS The nanosuspensions prepared by media milling method were solidificated by freeze drying method.With particle size and polydispersity index (PDI) as evaluation indices,stabilizer kind,ratio of diosgenin to stabilizer,ratio of preliminary nanosuspension volume to grinding bead amount,milling time,lyoprotectant kind and its amount as influencing factors,single factor test was applied to screening preparation and solidification processes.The morphology of nanosuspensions was observed,then the particle sizes and polydispersity indices of nanosuspensions and freeze-dried powder were determined.RESULTS The optimal conditions were determined to be 6:1 for ratio of diosgenin to Pluronic F127 (stabilizer Ⅰ),50:1 for ratio of diogenin to sodium dodecyl sulfate (SDS,stabilizer Ⅱ),1:4 for ratio of preliminary nanosuspension volume to grinding bead amount,120 min for milling time,and 8％ PEG-6000 and 2％ mannitol as lyoprotectants.The average particle size and polydispersity index of rod-like or flaky nanosuspensions were (348.1 ±14.2) nm and 0.244 ± 0.059,respectively,which were lower than those of freeze-dried powder.At room temperature,the particle sizes of nanosuspensions and freeze-dried powder remained stable within 35 d and 3 months,respectively.CONCLUSION The physical stability of diosgenin freeze-dried powder is superior to that of its nanosuspensions,which can be used after being reconstituted.

Although gene therapy was regarded as a promising approach for glioma treatment, its therapeutic efficacy was often disappointing because of the lack of efficient drug delivery systems. Mesenchymal stem cells(MSCs) have been reported to have a tropism for brain tumors and thus could be used as delivery vehicles for glioma therapy. Therefore, in this study, we attempted to treat glioma by using MSCs as a vehicle for delivering replication-competent adenovirus. We firstly compared the infectivity of type 3, type 5, and type 35 fiber-modified adenoviruses in MSCs. We also determined suitable adenovirus titer in vitro and then used this titer to analyze the ability of MSCs to deliver replication-competent adenovirus into glioma in vivo. Our results indicated that type 35 fiber-modified adenovirus showed higher infectivity than did naked type 3 or type 5 fiber-modified adenovirus. MSCs carrying replication-competent adenovirus significantly inhibited tumor growth in vivo compared with other control groups. In conclusion, MSCs are an effective vehicle that can successfully transport replication-competent adenovirus into glioma, making it a potential therapeutic strategy for treating malignant glioma.
Adenoviridae ; Animals ; Brain Neoplasms ; pathology ; therapy ; Cell Line, Tumor ; Genetic Vectors ; Glioma ; pathology ; therapy ; Humans ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation ; Oncolytic Virotherapy ; Random Allocation ; Virus Replication ; Xenograft Model Antitumor Assays

Medicinal animals are important part of Traditional Chinese medicine resources in China. Cytochrome c oxidase subunit I (COI) was selected as the standard DNA barcoding sequence for animal medical materials. In this study, the 51 animal species from 45 animal medical materials in the Chinese Pharmacopoeia were selected and the intra-specific variation and the inter-specific divergence, the barcoding gap, the identification efficiency of their COI sequences were analyzed. The results showed that the inter-specific divergence is higher than intra-specific distance. The barcoding gap existed between inter-specific sequence divergence and intra-specific dis-tance. The identification efficiencies were 100% both at the genus and species level except the Arthropoda. The cluster dendrogram exhibited that different species distinguished from others. Therefore, COI sequence as a bar-code is suitable to identify the species of animal medical materials in Chinese Pharmacopoeia.

Objective To explore the optimal experiment conditions of CCK-8 and MTS for cell proliferation assays in human amniotic epithelial cells and to evaluate the cytotoxicity of these reagents. Methods Human amniotic epithelial cells (hAECs) in logarithm growth stages were prepared in different cell concentrations with DMEM/F12 and 10% FBS. The sensitivity and optimal wavelengths was determined based on the optical density (OD) measured at 450 nm and 492 nm. The optimal time was determined under the conditions of the same cell concentration and defined OD values. HAECs were treated with DMSO, CCK-8 and MTS for 1 h, 2 h, 3 h, and 4 h, respectively. 24 h later, cytotoxicity of the CCK-8 and MTS was evaluated by determination of cell proliferation and Trypan Blue staining. Results The optimal detection wavelength was 450 nm for CCK-8, and 492 nm for MTS. The sensitivity of CCK-8 was slightly lower then that of MTS. The optimal time for incubation hAECs with CCK-8 was 4 h within 1~4 h. The inhibitory on cell proliferation and cytotoxicity of CCK-8 were weaker then those of MTS. Conclusion CCK-8 is a convenient reagent with low cytotoxicity for detection of the proliferation of hAECs.

Acquired Immunodeficiency Syndrome ; epidemiology ; transmission ; Adolescent ; Adult ; Anti-HIV Agents ; pharmacology ; China ; epidemiology ; Drug Resistance, Viral ; HIV-1 ; drug effects ; isolation & purification ; Humans ; Young Adult

OBJECTIVETo study the prevalence of primary HIV drug resistance in antiretroviral therapy (ART) areas of Henan province.

METHODSA total of 121 drug-naive long-term infected individuals and 154 patients with newly diagnosed from January 2011 to March 2012 were recruited, the questionnaires were surveyed and whole blood were collected to analyze the CD4(+)T cell counts and viral load. In-house method for genotypic resistance test was determined in those with viral load > 1000 copies/ml samples, the differences of demographic characteristics, immunological parameters and primary drug resistance were compared between the two groups.

RESULTSA total of 121 cases of long-term individuals who had infected (12.50 ± 3.21) years were mainly previous paid blood donors, and the age was (46.61 ± 9.32) years old. The infection route of the newly diagnosed were diversity, including blood, sexual transmission and others, the cases were 73, 73, 8, respectively, the confirmatory year was (0.91 ± 0.28) years, and average age was (22.21 ± 3.11) years old. The difference were statistically significant in the route of transmission, age and infection time from demographic analysis of the two groups (P < 0.05). The absolute M(P(25)-P(75)) counts of CD4(+)T lymphocytes of long-term group was 322 (217 - 422) cell/µl, which was lower than the newly diagnosed was 434(308 - 578) cell/µl (P < 0.05), and viral load was 4.0 (2.96 - 4.64) copies/ml, 3.77 (2.94 - 4.53) copies/ml, the difference was not significant (P > 0.05). The prevalence of primary drug resistance in long-term group and newly diagnosed was 5.79% (7/121), 9.09% (14/154), respectively, and the difference was statistically different (P < 0.05), and one PI-resistant strain was found in the newly diagnosed group.

CONCLUSIONThe primary drug resistant strains in untreated patients were found in Henan province of ART areas, and there was difference in degree of resistance between long-term infected individuals and newly diagnosed.

Adult ; Anti-HIV Agents ; pharmacology ; therapeutic use ; China ; epidemiology ; Drug Resistance, Viral ; Female ; HIV Infections ; drug therapy ; epidemiology ; virology ; Humans ; Male ; Middle Aged ; Viral Load

OBJECTIVETo explore the molecular mechanism and prevention of retinoic acid syndrome (RAS).

METHODSSDF-1 alpha mRNA from healthy adult lung tissue was measured by RT-PCR, CXCR4 protein expression on the cell membrane of APL cells induced by ATRA (APL-ATRA) was tested by FCM, and the rotary cell culture system (RCCS) was used to build a modal for in vitro stimulation of APL-ATRA infiltrating human lung tissue. The ability of APL-ATRA in adhesion, migration and infiltration was observed by interference from DEX, Ara-C and DNR.

RESULTSThe APL-ATRA cells could evidently infiltrate into normal lung tissue. Mean fluorescence intensity (MFI) of CXCR4 on the cell membrane of APL-ATRA cells was 30.6 +/- 1.8, which was much higher than that on unspecialized APL cells (9.8 +/- 4.2). SDF-1 alpha mRNA expression was detected positive in all 6 lung tissue. Contrary to the control groups, DEX could dramatically restrain the ability of APL-ATRA cells in adhesion and migration [(27.2 +/- 2.6)% vs. (46.0 +/- 3.0)%, (28.1 +/- 4.0)% vs. (48.2 +/- 3.0)%], while Ara-C and DNR could distinctly depress the ability in adhesion, migration and infiltration [(28.1 +/- 3.0)%, (30.2 +/- 3.2)% vs. (46.0 +/- 3.0)%; (29.0 +/- 4.0)%, (23.0 +/- 5.2)% vs. (48.2 +/- 3.0)%; (16.8 +/- 7.6)%, (17.1 +/- 6.0)% vs. (43.6 +/- 5.0)%].

CONCLUSIONIn vitro APL-ATRA cells can infiltrate into the human lung tissue. High expression of CXCR4 on APL-ATRA and SDF-1 alpha in the lung tissue may be one of the molecular mechanisms of the lung infiltration and RAS. DEX, Ara-C and DNR can dramatically restrain the ability of APL-ATRA cells in adhesion, migration and infiltration.

Adolescent ; Adult ; Cell Adhesion ; Cell Culture Techniques ; Cell Movement ; Chemokine CXCL12 ; genetics ; metabolism ; Child ; Female ; Humans ; Leukemia, Promyelocytic, Acute ; metabolism ; pathology ; Male ; Middle Aged ; Neoplasm Invasiveness ; Receptors, CXCR4 ; genetics ; metabolism ; Tretinoin ; adverse effects ; Tumor Cells, Cultured

Aim To investigate the effect of broneol on acute lung injury(ALI) induced by lipopolysaccharide (LPS). Methods Male C57 mice were randomly di-vided into saline group, model group, broneol group and dexamethasone group, then the ALI mouse model was induced by instilling intratracheally with LPS. The levels of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),interleukin-6(IL-6) and keratinocyte-de-rived cytokine (KC) were measured at 6h, 12h and 24h after instillation of LPS, and the pathological changes of lung were observed. Mice alveolar macro-phages (MHS) and epithelial cells (MLE-12) were stimulated by LPS. After the stimulation of 1h, 3h, 6h,9h, 12h, 24h, the levels of TNF-α and IL-6 in MHS cells and the contents of KC and macrophage in-flammatory protein-2 (MIP-2) in MLE-12 cells were measured. Results Broneol could inhibit the secre-tion of TNF-α,KC and IL-1β;the early effect of bro-neol on IL-6 was not obvious,but the later effect after the treatment of 24 hours was obvious. After LPS instil-lation 6h and 12h,Broneol could significantly improve lung tissue pathological changes. Broneol had no effect on TNF-α secretion of MHS cells, but it obviously af-fected IL-6 secretion in the later stage. In addition, broneol significantly inhibited KC and MIP2 secretion in MLE-12 cells at the later stage of LPS stimulation. Conclusions Broneol can protect LPS-induced acute lung injury. The mechanism may be related to the inhi-bition of the release of inflammatory factors,the activa-tion of inflammatory cells and the aggregation of neutro-phils.

Objective To clarify the genetic characteristics of human immunodeficiency virus (HIV) circulated in the population of men who have sex with men (MSM) in Zhengzhou,Henan and to analyze its relationship with HIV-1 prevailing in the paid blood donors (PBDs).Methods Thirty-one MSM who were confirmed as HIV positive individuals in 2010 together with 41 HIV-positive former PBDs were enrolled in the study.Information on related epidemiological characteristics and their plasma were collected.RT-PCR was used to amplify HIV-1 full length gag (1584 bp),pol (3147 bp) genes and partial env gene (C2V3 segment,558 bp) followed by sequencing on those subjects.Online software available at LosAlamos HIV Database was used to identify the HIV subtypes based on the findings of the sequences.Phylogenetic tree was used to identify thc possible relationship of transmission.Results Fifty-three full length gag,38 full length pol and 48 partial env (C2V3) genes were collected from 72 participants.Among the 31 HIV ( + )MSM individuals,14 CRF01_AE strains,5 CRF07BC_ strains and 12 subiype B ( 1 subtype B and 11B' ) strains were identified respectively.All of the 41 strains identified from former PBDs were infected by B' strains.The CRF01_AE strains identified in MSM showed a close relationship to those identified from both Hebei and Liaoning provinces.The CRF07 BC strains showed a close relationship with those from Shijiazhuang and Beijing cities.Among the 12 subtype B strains,8 sequences grouped into 1 cluster with 1 sequence from the former PBDs.Two sequences grouped with 02HNseq4 suggested that B' had been prevailed in the MSM population might come from the former PBDs and were closely related to the strains identified in the MSM population.Conclusion Complicated genetic background and multiple introductions of HIV in the MS population in Zhengzhou,were found.This was also the first report which noticed that the subtype B epidemic among Zhengzhou MSM was mainly originated from the B' among the former PBDs.

Objective To analyze the situation of AIDS patients who had received replaced therapy program in Henan province.Methods 84 AIDS patients had been enrolled into the national free first-line antiretroviral treatment for more than 5 years and would soon be replaced with another antiretroviral treatment program,were selected to a follow-up program to be carried out six months later Data on CD4+ tlymphocyte count,viral load and genotypic resistance were included in the study.Results The DDI + AZT + NVP treatment program was used by all the 84 patients at baseline.A replacement by 3TC + AZT + NVP (post first-line) in 31 patients and 3TC + TDF + LPV/r (second-line) in another 53 patients were taken place within a week.All the patients were followed for six months.Results showed that:all of the 84 patients appeared an amelioration of CD4 1 tlymphocyte count median from the baseline of 374.00 cell/μl to 406.50 cell/μl(P=0.005).Those patients who had changed to second-line treatment program also showed an improvement of CD4 + tlymphocyte count median from the baseline of 267.00 cell/μl to 365.00 cell/μl (P=0.015),while patients who were on the post first-line program with their CD4 + tlymphocyte count mean did not show significant change as compared to the baseline (P=0.158) data.All the 84 patients showed a decrease of virus load median from the baseline of 3.61 Iog10copies/ml to 0.00 log10 copies/ml (P=0.000).Both of the two types of patients who had been changed to different programs,had a lower virus load median in the end of the tollow-up period (for post first-line:P=0.007 ；for second-line:P=0.000).13 patients kept their viral load more than 1000 copies/ml,including 5 cases bore more than three thymidine analogue mutations (TAMs) a the end of the follow-up program.Another 4 patients had no resistance mutations detected and no significant variation of viral load (less than 3 times) in the pre-or post- surveys.Conclusion AIDS patients who had received long-term first-line antiretroviral treatment program,showed an amelioration six months after changing of the treatment program.Timely and effective testing on drug resistance as well as the strengthening of the follow-up program still seemed to be the link to those patients who were receiving first-line treatment that should not be ignored.