OBJECTIVETo explore the effect of Xinfeng Capsule (XC) on lipoprotein metabolism of rheumatoid arthritis (RA) patients.

METHODSTotally 180 RA patients were assigned to the experimental group and the control group by random digit table, 90 in each group. Patients in the experimental group took XC (three pills each time, three times daily), while those in the control group took Methotrexate Tablet (four tablets each time, once per week). One month consisted of one therapeutic course and all patients were treated for two therapeutic courses. A healthy control group consisting of 60 patients was also set up. Changes of lipoprotein indices, clinical efficacy, lipid metabolism, joint symptoms and signs, activity indicators were observed, and correlation analyses were performed.

RESULTSCompared with the healthy control group, expression levels of prealbumin (PA), globulin (GLO), high-density lipoprotein (HDL), apolipoprotein Al (Apo-A1) were lowered in RA patients (P <0. 05, P <0. 01). Correlation analyses showed that PA was negatively correlated with joint tenderness, morning stiffness time, disease activity score (DAS-28), C-reactive protein (CRP), interleukin (IL)-6, respectively. Total protein (TP) was negatively correlated with joint tenderness. GLO was negatively correlated with joint tenderness and DAS-28. HDL was negatively correlated with erythrocyte sedimentation rate (ESR) and endothelin (ET)-1. Apo-Al was negatively correlated with joint pain; Apo-B was negatively correlated with CRP; LDL was negatively correlated with morning stiffness time (P <0. 05, P <0. 01). Compared with before treatment, expression levels of PA, HDL, Apo-A1 , Apo-B, and serum IL-10 contents increased, and expression levels of ESR, CRP, IL-6, ET-1 , joint pain, joint swelling, morning stiffness time, and DAS-28 decreased in the experimental group (P <0. 05, P <0. 01). PA increased more after treatment than before treatment in the control group (P <0. 01). There was statistical difference in joint symptoms (except joint tenderness) and activity indices (except ET-1) in the control group (P <0. 05, P <0. 01). Compared with the control group after treatment, PA and HDL increased, ET-1 and duration of morning stiffness decreased in the experimental group (all P <0. 05).

CONCLUSIONSLipoprotein metabolic disorder exists in RA patients, and it is associated with disease activity. XC could obviously improve lipoprotein metabolism and joint symptoms.

Arthritis, Rheumatoid ; drug therapy ; metabolism ; Blood Sedimentation ; C-Reactive Protein ; Capsules ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Humans ; Interleukin-10 ; Interleukin-6 ; Lipoproteins ; Lipoproteins, HDL ; metabolism ; Methotrexate

Candida shehatae xyl1 gene and Pichia stipitis xyl2 gene were amplified by PCR and the xyl1 and xyl2 were both placed under the promoter GAL of vector pYES2 to produce the recombinant expression vector pYES2-P12. Subsequently the pYES2-P12 vector was transformed into S. cerevisiae YS58 by LiAc to produce the recombinant yeast YSS8-12. It was indicate that the recombinant yeast YSS8-12 could converse xylose to ethanol with the xylose consumption rate of 81. 3%.

The specimens of ductal carcinoma in situ (DCIS) with early invasion, and specimens collected by core needle biopsy (CNB) tend to contain limited amount of invasive component, so it is imperative to explore a new technique which can assess HER2 gene status accurately for the limited invasive cancer component in these specimens. Dual staining technique of combining immunohistochemistry (IHC) for myoepithelial cells and single or dual probe chromogenic in situ hybridization (CISH) for HER2 gene was performed on routinely processed paraffin sections from 20 cases diagnosed as having DCIS with invasive cancer. Among them, 10 had fluorescence in situ hybridization (FISH)-confirmed amplification of HER2 and 10 had FISH-confirmed non-amplification of HER2. We successfully detected HER2 genetic signals and myoepithelial IHC markers (SMM-HC or CK5/6) simultaneously on a single section in all 20 specimens. Myoepithelial markers and HER2 signals detected by dual staining assay were consistent with those by individual technique performed alone. HER2 gene amplification results determined by dual staining assay were 100% consistent with those of FISH. Dual staining technique which allows simultaneous detection of myoepithelial marker protein and cancerous HER2 gene is feasible, and it has potential to be used in clinical practice for effective determination of HER2 amplification in limited invasive component.
Biomarkers, Tumor ; metabolism ; Breast Neoplasms ; genetics ; metabolism ; pathology ; Chromogenic Compounds ; Female ; Gene Expression Profiling ; methods ; Humans ; Immunohistochemistry ; methods ; In Situ Hybridization, Fluorescence ; methods ; Neoplasm Invasiveness ; pathology ; physiopathology ; Receptor, ErbB-2 ; genetics ; metabolism ; Reproducibility of Results ; Sensitivity and Specificity

OBJECTIVETo explore the killing effect of the mutant herpes simplex virus thymidine kinase (HSV-sr39tk) and its wild-type (HSV-tk) mediated by lentiviral vector on T lymphocytes in vitro and compare T cell survival rate after GCV or ACV treatment.

METHODSThe three-plasmid lentiviral vector system including packaging plasmid DeltaNRF, envelope plasmid VSV-G and vector plasmid (pTK151 + HSV-sr39tk or pTK151 + HSV-tk) were cotransfected into human embryonic kidney 293T cells using modified calcium phosphate precipitation methods. The packaged virus was harvested 72 h later. The survival of T cells expressing HSV-sr39tk or HSV-tk was measured by MTT assay after 4 day-culture against a gradient of GCV or ACV concentrations.

RESULTSThe three plasmids were effectively cotransfected and a high titre of lentivirus was obtained (2 x 10(6) IU/ml). 39tk(+) T cell survival rates declined promptly when the prodrug GCV/ACV concentrations increased from 0 micromol/L to 10 micromol/L. The T cell survival rates in GCV group declined from (96.04 +/- 3.23)% to (36.76 +/- 4.38)% while in ACV group from (97.31 +/- 4.61)% to (43.75 +/- 8.99)%. However, when GCV/ACV concentrations were more than 10 micromol/L, further decline of 39tk(+) T cell survival rates became unobvious. The growth rate of 39tk(+) T cell exposed to GCV or ACV was obviously lower than that in un-transfected T cells (P < 0.05). Tk(+) T cells were sensitive to GCV (P < 0.05), but not to ACV (P > 0.05). There was a significant difference in killing effects between 39tk(+) T cell + GCV group and tk(+) T cell + GCV group (P < 0.05).

CONCLUSIONThe lentiviral vectors containing HSV-sr39tk gene could infect T lymphocytes effectively and stably without affecting the proliferation of the transduced cell. In contrast to HSV-tk gene, T cells infected HSV-sr39tk were more sensitive not only to GCV but also to ACV.

Acyclovir ; pharmacology ; Animals ; Cell Survival ; drug effects ; Cells, Cultured ; Ganciclovir ; pharmacology ; Genetic Vectors ; Lentivirus ; genetics ; Mice ; Mice, Inbred C57BL ; Plasmids ; genetics ; T-Lymphocytes ; cytology ; drug effects ; Thymidine Kinase ; genetics ; Transfection

OBJECTIVETo explore the expression of beta protein 1 (BP1) gene in adult acute leukemia (AL) and its relationship with acute leukemia.

METHODSExpression of BP1 gene mRNA was detected in 70 adult AL, 10 normal controls and HEL cell line, by reverse transcriptase-polymerase chain reaction (RT-PCR).

RESULTSNo detectable BP1 gene was found in peripheral blood or bone marrow cells of normal controls and 20 acute myeloid leukemia (AML) in complete remission (CR) stage. BP1 gene was highly expressed in HEL cell line, 57% (20/35) of AML and 80% (8/10) of AML-M(5) cases. BP1 gene could not be detected in adult acute lymphoid leukemia.

CONCLUSIONBP1 gene was highly expressed in AML. It might be used as a molecular marker of AML.

Acute Disease ; Adolescent ; Adult ; Female ; Genes, Homeobox ; Globins ; genetics ; Homeodomain Proteins ; genetics ; Humans ; Leukemia ; genetics ; Male ; Middle Aged ; Transcription Factors ; genetics

This study was aimed to propagate and identify the prdm1 gene-knockout mice, so as to lay the foundation for studying Blimp-1 protein. Two kinds of transgenic homozygous mice with B6.prdm1(flox/flox) and B6.Lck-Cre were feed and propagated; after successful propagating, the first passage mice were obtained; after the first passage mice were copulated once again, the genotypes were obtained as follows: B6. prdm1(wild/wild). Lck-Cre, B6. prdm1(wild/wild), B6.prdm1(flox/flox). Lck-Cre, B6.prdm1(flox/wild). Lck-Cre, B6.prdm1(flox/flox), B6. prdm1(flox/wild). The genomic DNA of second passage mice was extracted, the Cre and loxp gene fragments were amplified by PCR, then the size of Cre and loxp genomic DNA were detected by agarose gel electrophoresis. The mice with B6.prdm1(flow/flox). Lek-Cre were used as conditionally prdm1-knockout mice, B6.prdm1(flox/wild). Lck-Cre mice, B6.prdm1(flox/flox) and B6 mice were used as controls. The spleen T lymphocytes and B lymphocytes were sorted by using magnetic beads, the blimp-1 target protein was identified by Western blot. The results showed that the two transgenic homozygous mice had the ability to reproduce, and the separation ratio of second passage mice generated from propagation of their offspring cach other meet Mendelian laws, and the prdm1 gene-knockout mice also could successfully obtained. It is concluded that the application of Cre-loxp system may successfully obtain plentiful prdm1 gene-knockout mice.
Animals ; Genotype ; Mice ; Mice, Inbred C57BL ; genetics ; Mice, Knockout ; genetics ; Reproduction ; Transcription Factors ; genetics

OBJECTIVETo study the plasma protein binding rate of methyl protodioscin.

METHODThe ultrafiltration was employed to determine the plasma protein binding rate of methyl protodioscin. The plasma concentrations of methyl protodioscin were measured by HPLC-MS-MS.

RESULTThe plasma protein binding rate of methyl protodioscin with rat plasma at the concentration of 20.0, 100 and 200 microg x mL(-1) were (94.6 +/- 0.16)%, (91.6 +/- 0.35)% and (86.10 +/- 0.60)%, respectively, while the plasma protein binding rate of methyl protodioscin with normal human plasma at the above concentrations were (82.11 +/- 5.12)%, (84.54 +/- 0.32)% and (88.52 +/- 1.02)%, respectively.

CONCLUSIONThe binding rate of methyl protodioscin with plasma protein is high.

Animals ; Antineoplastic Agents ; metabolism ; Blood Proteins ; metabolism ; Calibration ; Chromatography, High Pressure Liquid ; Diosgenin ; analogs & derivatives ; metabolism ; Female ; Humans ; Male ; Protein Binding ; Rats ; Saponins ; metabolism ; Sensitivity and Specificity ; Tandem Mass Spectrometry ; Ultrafiltration

This study was aimed to investigate the protective effect of Wit3a gene modification on mouse bone marrow mesenchymal stem cells against the injury induced by Ara-C. The gene-modified MSC steadily expressing Wnt3a were established by adenovirus system. The acute direct damage effects of different concentrations of Ara-C on the unmodified MSC and the gene-modified MSC were assessed by using an in vitro culture system, and the corresponding controls were set. The proliferation and apoptosis of MSC exposed to Ara-C were detected by cell count kit-8 (CCK-8) and flow cytometry. The expression of BCL-2 protein related with cell apoptosis was assayed by Western blot. The results indicated that as compared with unmodified MSC, Ara-C exhibited a less inhibitory effect on the proliferation of gene-modified MSC. There was obvious difference between unmodified MSC and gene-modified MSC (p < 0.05). The proliferation of gene-modified MSC began to recover at 72 hours after removal of Ara-C. However, unmodified MSC showed sustained suppression of proliferation after withdrawal of Ara-C. In apoptosis, the apoptosis rate of gene-modified MSC induced by Ara-C was significantly lower than those of unmodified MSC (p < 0.05). In addition, the expression levels of BCL-2 protein in gene-modified MSC were up-regulated compared with unmodified MSC (p < 0.05). It is concluded that Wnt3a gene modification can significantly mitigate the damage of mouse bone marrow MSC induced by Ara-C.
Animals ; Bone Marrow Cells ; drug effects ; metabolism ; Cytarabine ; adverse effects ; Mesenchymal Stromal Cells ; drug effects ; metabolism ; Mice ; Organisms, Genetically Modified ; Proto-Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; Wnt3A Protein ; genetics

OBJECTIVETo establish strategies for preventing graft versus host disease (GVHD) and reducing treatment associated morbidity while preserving graft versus leukemia (GVL) effect in nonmyeloablative allogeneic bone marrow transplantation (allo-BMT), with or without donor lymphocyte infusion (DLI) after BMT.

METHODS3 x 10(7) bone marrow cells mixed with 1 x 10(7) spleen cells from the same BALB/c mouse were transplanted into the nonablative irradiated inbred 615 mouse which received a single subcutaneous injection of 1 x 10(6) L615 leukemia cells three days before. The experiments were designed as follows (ten mice in each group): myeloablative BMT control group (group A), nonmyeloablative conditioning without BMT group (group B), nonmyeloablative BMT group (group C), and nonmyeloablative BMT + DLI group (group D). GVL effects were assessed by survival time, white blood cell count and L615 cells in peripheral blood and histologic changes. GVHD was assessed by signs of weight loss, ruffled fur, diarrhea and histologic changes of skin, liver and small intestines. Chimerism was detected by cytogenetic analysis and PCR technique.

RESULTSThe survival time of group A, B, C and D was (20.3 +/- 13.4), (15.9 +/- 1.1), (21.6 +/- 1.7) and (37.8 +/- 2.0) days, respectively, being no significant difference between group A and group C (P > 0.05). The survival time of group C was longer than that of group B (P < 0.01). And among group B, C and D, group D had the longest survival time (P < 0.01). GVHD signs and histologic changes were observed in 60% of control group mice at + 14 day, but none of group C and group D. 40% of mice in group A died of treatment associated morbidity within two weeks, but none in group C and group D. Allogeneic chimerism was kept in group A, but excluded gradually in group C.

CONCLUSIONGVL effect seems preserved in nonmyeloablative BMT mice, but weaker than that in myeloablative BMT mice. GVL effect seems to be enhanced by DLI after nonmyeloablative BMT. GVHD and transplantation associated morbidity seems to be reduced in nonmyeloablative BMT.

Animals ; Bone Marrow Transplantation ; immunology ; methods ; Combined Modality Therapy ; Female ; Graft vs Host Disease ; prevention & control ; Graft vs Leukemia Effect ; Leukemia, Experimental ; therapy ; Leukemia, Lymphoid ; therapy ; Lymphocyte Transfusion ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Inbred Strains ; Transplantation Conditioning ; methods ; Transplantation, Heterologous

OBJECTIVETo explore the influence of the lentiviral vector mediated mouse genetic engineering regulatory T cells (Treg) infusion on post-allogeneic bone marrow transplantation (allo-BMT) acute graft-versus-host disease (GVHD) in mice.

METHODSLentivirus-mediated Forkhead box P3 (Foxp3) gene was transduced into BALB/c mice CD4(+)CD25(-) T cells (Treg) to construct engineered Tregs in vitro. An allo-BMT model of BALB/c to C57BL/6 mice was established. After irradiation, the recipients were injected with donor cells plus the genetic engineering Tregs. Survival time, clinical GVHD score, histopathological findings, activation of donor T cells or serum levels of inflammatory cytokines were observed after allo-BMT.

RESULTSThe mean survival times for radiation alone group (Gp I), transplantation control group (Gp II), engineering Treg infusion group (Gp III) and empty vector control group (Gp IV) were (8.8 ± 0.6) d, (36.7 ± 2.5) d, (51.6 ± 4.0) d and (34.1 ± 2.3) d, respectively. The survival time was significantly longer in Gp III than in other groups (P < 0.05). Histopathological finding in several target organs (skin, liver and small intestine) confirmed the presence of severe GVHD in Gp II and Gp IV, while no histological signs of GVHD were observed in long survival recipients in Gp III, and clinical GVHD scores in Gp III were significantly lower than that in Gps II and IV. The numbers of donor T cells and the percentage of IFN-producing donor T cells in the spleen of recipients in Gp III were significant lower than those in Gps II and IV at days 3 and 4, and at day 3 after transplantation, respectively (P < 0.05). The serum levels of IFN-γ, IL-2 and TNF-α were increased at day 21 to 28 after transplantation in all groups. The peak concentrations of IFN-γ, IL-2 and TNF-α in Gp III were significantly lower than those in Gps II and IV control groups at day 21 (P < 0.05).

CONCLUSIONCo-injection of genetic engineering Treg can efficiently prevent recipients from lethal GVHD after allo-BMT in mice by inhibiting the early activation and expansion of donor T cells and reducing the serum levels of inflammatory cytokines.

Animals ; Bone Marrow Transplantation ; adverse effects ; Cytokines ; blood ; Female ; Forkhead Transcription Factors ; genetics ; Genetic Engineering ; Genetic Vectors ; Graft vs Host Disease ; immunology ; Lentivirus ; Lymphocyte Activation ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; T-Lymphocytes, Regulatory ; cytology ; immunology ; Transduction, Genetic ; Transplantation, Homologous