OBJECTIVETo investigate the regulation pattern of the two compound prescriptions for Kidney tonifying, Yougui Yin and Bushen Yishou capsule, in down-regulating T-cell apoptosis gene expression in aged rats.

METHODSExpressions of T-cell apoptosis promoting and inhibiting genes, including Fas, FasL, Bcl-2, Bax, TNFR1 and TNFR2, as well as activity of cysteine proteinase in cascade connection, such as Caspase 8 and Caspase 3 were determined by TUNEL labeled flow cytometry and fluorescence real-time quantitative RT-PCR technique. The difference between old and young rats was compared, and the different regulation patterns of the two compound prescriptions for Kidney tonifying and their effects on Caspase activity were compared with those of compound prescription for blood circulation activating.

RESULTSThe two compound prescriptions for Kidney tonifying could effectively lower T-cell over-apoptosis in old rats, down-regulate FasL and TNFR1 gene transcription, and decrease the activity of Caspase 8 and Caspase 3, while the compound prescription for blood circulation activating showed insignificant effect on T-cell over-apoptosis.

CONCLUSIONKidney-deficiency is closely related to the T-cell over-apoptosis. The T-cell over-apoptosis in old rats could be effectively improved by the two compound prescription for Kidney tonifying through down-regulating the apoptosis promoting genes FasL and TN-FR1 transcription. That is the unique regulation pattern of Kidney tonifying principle to T-cell apoptosis related gene in old rats.

Aging ; drug effects ; genetics ; immunology ; Animals ; Apoptosis ; drug effects ; Blood Circulation ; physiology ; Caspase 3 ; Caspases ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Fas Ligand Protein ; Female ; Male ; Membrane Glycoproteins ; biosynthesis ; genetics ; Random Allocation ; Rats ; Receptors, Tumor Necrosis Factor ; biosynthesis ; genetics ; T-Lymphocytes ; cytology ; Yang Deficiency ; immunology ; fas Receptor ; biosynthesis ; genetics

OBJECTIVETo observe the clinical curative effect of Kidney tonifying method on retardation of immunosenescence and corresponding experimental study.

METHODSA randomized double blind placebo-controlled trial was used (RCT) on 22 pairs of aged subjects to elucidate the effect of Kidney tonifying recipe on the peripheral T-lymphocyte apoptosis and the Fas/FasL gene expression in them. In rats experimental study, the effects of two kinds of Chinese recipes (Kidney tonifying recipe and blood circulation promoting recipe) on the same parameters as in clinical study as well as on cell apoptosis and gene expression regulation in old rats were also observed.

RESULTSClinical study showed that after treatment, the percentage of T-lymphocyte apoptosis and the FasL gene expression in the Kidney tonifying group of aged subjects were lower than those in the placebo group (P < 0.01). Animal experiment showed the same result as shown in clinical study in Kidney tonifying recipe treated rats, but not shown in those treated with blood circulation promoting recipe statistically.

CONCLUSIONKidney tonifying principle has down-regulating effect on the transcription of apoptotic gene in both aged persons and old rats, this is one of the molecular mechanisms of Kidney tonifying method in decreasing over-apoptosis in aged subjects and old rats.

Aged ; Aged, 80 and over ; Aging ; drug effects ; Animals ; Apoptosis ; drug effects ; Double-Blind Method ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Fas Ligand Protein ; Female ; Humans ; Male ; Medicine, Chinese Traditional ; Membrane Glycoproteins ; biosynthesis ; genetics ; Middle Aged ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Sprague-Dawley ; T-Lymphocytes ; cytology ; fas Receptor ; biosynthesis ; genetics

OBJECTIVETo investigate the mechanism of senescence delay of human diploid fibroblast (2BS) and protecting telomere length by epimedium flavonoids (EF).

METHODSThe drug sera of EF were used to treat the 2BS. The population doublings of 2BS cells were observed, the mRNA expression of p16 gene were determined by fluorescence real-time quantitative RT-PCR, the telomerase activation of 2BS cells were determined by TRAP-Hyb, the total retinoblastoma (Rb) and phosphorated Rb protein content were detected by ELISA, the telomere length of 2BS cells were determined by telomere restriction fragment (TRF) Southern blot assay.

RESULTSEF could significantly extend the population doublings of 2BS cells, the expression of p16 mRNA was decreased and the content of phosphorated Rb protein were increased by EF. The telomere lengthening of 2BS cells were improved by EF, but the telomerase was not activated.

CONCLUSIONIn senescence human fibroblasts 2BS cells, p16 gene mRNA expression increased, content of phosphorated Rb protein decreased and the telomere length of 2BS shortened, EF might delay the aging of cells through inhibiting the p16 gene expression, promoting the production of phosphorated Rb protein and to protect the length of telomere, but not activating the telomerase.

Animals ; Cells, Cultured ; Cellular Senescence ; genetics ; Cyclin-Dependent Kinase Inhibitor p16 ; biosynthesis ; genetics ; Epimedium ; chemistry ; Fibroblasts ; cytology ; Flavonoids ; pharmacology ; Male ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Sprague-Dawley ; Retinoblastoma Protein ; metabolism ; Telomerase ; biosynthesis ; Telomere ; genetics ; metabolism ; Transduction, Genetic

OBJECTIVETo elucidate the gene regulatory pattern of Epimedium flavonoids (EF) in immune homeostasis remodeling in the aged rats.

METHODS(1) To quantitatively analyse the apoptosis percentage of lymphocyte in spleen of aged, young and EF treated rats using flow cytometry. (2) To analyse the lymphocyte gene expression profiles of different groups using gene chips (Rat Genome U34A).

RESULTS(1) Detection of lymphocyte apoptosis percentage showed that there was significant difference in comparing between aged group and young group, between EF treated group and aged group (P < 0.01). (2) As compared with that in the young group, in the aged group, 116 genes were up-regulated and 215 down-regulated. As compared with that in the old group, in the EF treated group, 447 genes were up-regulated and 456 down-regulated, which involved the aspects as cell apoptosis and cell proliferation regulation, etc.

CONCLUSION(1) The expression pattern characterized by up-regulation of apoptosis promoting genes expression and down-regulation of apoptosis inhibiting genes expression, is the important gene background of immuno-homeostasis imbalance in the aged. (2) The role of EF is to reverse the abnormal changes of gene expressions with opposite functions, i.e. the apoptosis promoting and inhibiting, proliferation enhancing and antagonizing genes, to reconstruct a beneficial equilibrium of gene expression and thus to further remodel the immuno-homeostasis in the aged.

Adjuvants, Immunologic ; pharmacology ; Aging ; Animals ; Apoptosis ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Epimedium ; chemistry ; Female ; Flavonoids ; pharmacology ; Gene Expression ; Gene Expression Profiling ; Homeostasis ; Lymphocytes ; immunology ; metabolism ; Male ; Neuroimmunomodulation ; drug effects ; Rats ; Rats, Sprague-Dawley ; T-Lymphocytes ; immunology ; metabolism

AIM To establish an HPLC method for the simultaneous content determination of echinacoside,verbascoside,aucubin,pinoresinol diglucoside,epimedin A,epimedin B,epimedin C,icariin and baohuoside Ⅰ in Jingang Tablets (Cistanches Herba,Eucommiae Cortex,Epimedii Folium,etc.).METHODS The analysis of 50％ methanol extract of this drug was performed on a 30 ℃ thermostatic Agilent TC-C18 column (250 mm × 4.6 mm,5 μm),with the mobile phase comprising of (methanol-acetonitrile)-0.2％ phosphoric acid flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelengths were set at 330,208 and 270 nm.RESULTS Nine constituents showed good linear relationships within their own ranges (r ＞ 0.999 0),whose average recoveries were 96.64％-100.08％ with the RSDs of 0.71％-1.76％.CONCLUSION This sensitive and accurate method can be used for the quality control of Jingang Tablets.

UNLABELLEDTo investigate the regulatory effects of epimedium flavonoids (EF) on adrenocortical regeneration in rats with inhibited hypothalamic-pituitary-adrenal (HPA) axis.

METHODSCell distribution in cell cycle and cell apoptotic rate were measured with PI stain and flow-cytometry; apoptosis cells were showed by in situ terminal-deoxynucleotidyl transferase-mediated deoxy-uridine triphosphate-fluorescene nick end labeling assay (TUNEL), and the genome-wide gene mRNA expression was detected by oligonucleotide microarrays.

RESULTSCompared to the normal control, adrenal cells isolated from the HPA axis inhibited model group were arrested in Go/GI phase, and showed a higher apoptotic rate (P < 0.05). After treated with EF, cells in G0/G1 phase decreased and those in G2/M phase increased (P < 0.01), and the elevated apoptotic rate reduced significantly (P < 0.05). TUNEL assay showed the number of apoptotic cells per section was 4.67 1.53 in the normal control group, 70.67 +/- 9.29 in the model group, and 18.67 +/- 7.64 in the EF-treated group respectively (n=3). Gene expressions in adrenal were mostly restrained in the model group, including 7 cytocycle promoting genes, including V-ras, V-jun, etc., while after treatment with EF, 6 cytocycle promoting genes, 1 anti-apoptotic gene, and genes that closely related with adrenocortical regeneration as IGF-II and FGF7 and their receptors, as well as 7 steroid biosynthesis participated genes were all up-regulated. Conclusion EF can accelerate adrenocortical cell proliferation, inhibit its apoptosis, and promote steroid biosynthesis so as to enhance adrenocortical regeneration in HPA axis inhibited rats, which may contribute to the beneficial effects of EF in protecting adrenocortical function during glucocorticoid withdrawal.

Animals ; Drugs, Chinese Herbal ; pharmacology ; Epimedium ; chemistry ; Flavonoids ; pharmacology ; Gene Expression ; Gene Expression Profiling ; Glucocorticoids ; adverse effects ; Hypothalamo-Hypophyseal System ; drug effects ; Male ; Oligonucleotide Array Sequence Analysis ; Pituitary-Adrenal System ; drug effects ; physiology ; Rats ; Rats, Sprague-Dawley ; Substance Withdrawal Syndrome ; prevention & control

OBJECTIVETo compare the modulating effect of Kidney tonifying recipe (KTR) and Spleen invigorating recipe (SIR) on T-lymphocyte apoptosis (TLA) in corticosterone treated rats (CTR).

METHODSQualitative and quantitative analysis on activation-induced TLA was conducted by transmission electron microscope and TUNEL labelled flow cytometry detection.

RESULTSTLA percentage of CTR was 45.87 +/- 7.22%, while that of normal control rats was 34.25 +/- 6.47%, the difference between them was significant (P < 0.01). TLA percentage of Youguiyin treated rats was 35.90 +/- 7.39%, and that of Bushen Yishou capsule treated rats was 36.20 +/- 9.14%, compared with that of CTR, P < 0.01 and P < 0.05 respectively. TLA percentage of the SIR treated group was 36.92 +/- 11.82%, which was different insignificantly as compared with that of CTR.

CONCLUSIONTLA susceptibility was significantly enhanced in CTR. Both KTRs (Youguiyin and Bushen Yishou capsule) had down-regulating effect on TLA, while the effect of SIR was insignificant, suggesting that down-regulating activation-induced TLA may be one of the important mechanisms of KTR in improving T-lymphocyte function in CTR.

Animals ; Apoptosis ; drug effects ; Corticosterone ; Drugs, Chinese Herbal ; pharmacology ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; T-Lymphocytes ; cytology ; ultrastructure ; Yang Deficiency ; chemically induced ; immunology

OBJECTIVETo investigate the clinical effect of bridging fixation with locking plate for the Seinsheimer type V subtrochanteric femoral fracture.

METHODSFrom March 2009 to September 2014,18 cases of Seinsheimer type V subtrochanteric femoral fracture were treated by open reduction and bridging fixation with locking plate through proximal and distal approach including 16 males and 2 females with an average age of 41 years old ranging from 22 to 67 years old. Among them, 12 cases caused by traffic accident, 5 cases by falling, 1 case by heavy aboving. All cases were fresh and closed fractures. Time between injury and operation was from 4 to 9 days with an average of 6.2 days. Of them, 11 cases were fixed with reverse LISS and the other 7 cases were fixed with anatomical locking plates of proximal femur.

RESULTSThe mean time of operation was 110 min (ranged from 90 to 155 min). The mean blood loss during operation was 425 ml (ranged from 350 to 650 ml) and 16 cases got blood transfusion which was meanly 300 ml. The mean hospital time was 14 days (ranged from 12 to 18 days). The mean duration of followed up was 11.8 months (ranged from 8 to 22 months). The mean time of bone union was 6.6 months (ranged from 5 to 8 months). There was not any complication such as infection, implant failure, hip varus, external rotation deformity of low limb or fat embolism. The Sanders hip scores were 53.22 ± 6.48, the result was excellent in 12 cases and good in 6 cases at the last follow-up.

CONCLUSIONUnder the principle of biological osteosynthesis, treatment of Seinsheimer type V subtrochanteric femoral fracture with bridging locking plate fixation has such advantages as high mechanism, less interference of blood supply, stable fixation and little complication. It is a safe and idea way for the treatment of the Seinsheimer type V subtrochanteric femoral fracture.

Adult ; Aged ; Bone Plates ; Female ; Fracture Fixation, Internal ; methods ; Hip Fractures ; surgery ; Humans ; Male ; Middle Aged

OBJECTIVETo investigate the effects of tributyrin (TB), a histone deacetylase inhibitor, on the growth, differentiation and apoptosis of SHI-1 leukemia cells and explore its possible mechanism.

METHODCell proliferation and viability were determined by cell counting, trypan blue dye exclusion. Cell morphological analysis, Annexin binding, DNA electrophoresis, expression of CD11b and CD14, NBT reduction were performed to evaluate differentiation and apoptosis of SHI-1 cells. The level of acetylated histone H3 was detected by Western blot and p21(WAF1/CIP1) expression by semi-quantitative RT-PCR.

RESULTSTB inhibited the proliferation and viability of SHI-1 cells in a time-dose dependent manner. The morphology of SHI-1 cells cultured in the presence of 0.1 mmol/L TB for 72 hs was more mature with higher NBT positivity and up-regulated expressions of CD11b and CD14 than that of control group. Exposed to 0.5 - 1.0 mmol/L TB for 48 hs, SHI-1 cells exhibited the morphological hallmarks of apoptosis, the increasing of Annexin binding and the DNA ladder on gel electrophoresis. The level of acetylated histone H3 and p21(WAF1) mRNA extracted from SHI-1 cells were increased by the treatment of TB.

CONCLUSIONTB can inhibit proliferation, induce differentiation and apoptosis of SHI-1 cells. The mechanism may associate with its up-regulation of acetylated histone and the expression of p21(WAF1).

Acetylation ; drug effects ; Apoptosis ; drug effects ; Blotting, Western ; Cell Differentiation ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cyclin-Dependent Kinase Inhibitor p21 ; genetics ; Enzyme Inhibitors ; pharmacology ; Gene Expression ; drug effects ; Histone Deacetylase Inhibitors ; Histone Deacetylases ; metabolism ; Histones ; metabolism ; Humans ; Leukemia, Monocytic, Acute ; genetics ; metabolism ; pathology ; Reverse Transcriptase Polymerase Chain Reaction ; Triglycerides ; pharmacology

To elucidate the possible mechanism of differentiation and/or apoptosis in NB4, K562 cells induced by tributyrin (TB), a histone deacetylase inhibitor (HDACi), the level of acetylated histone H3 was detected by Western blot, p21(WAF1) expression was detected by semi-quantitative RT-PCR. The results showed that histone H3 hyperacetylation was detected in NB4 (or K562) cells after treatment with TB 0.1 mmol/L (or TB 0.5 mmol/L) for 16 hours in a dose-dependent manner. p21(WAF1) dose-dependently increased at RNA level in these two cell lines treated by TB 0.1 mmol/L. The level of p21(WAF1) mRNA increased at 2 hours after TB treatment, reaching peak at 16 hours, and maintaining for 48 hours. In conclusion, the mechanism of differentiation and apoptosis in NB4, K562 cells induced by tributyrin may associate with its up-regulation of acetylated histone and p21(WAF1) mRNA level.
Acetylation ; drug effects ; Apoptosis ; drug effects ; Blotting, Western ; Cell Differentiation ; drug effects ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p21 ; genetics ; Dose-Response Relationship, Drug ; Enzyme Inhibitors ; pharmacology ; Gene Expression Regulation, Neoplastic ; drug effects ; Histone Deacetylase Inhibitors ; Histone Deacetylases ; metabolism ; Histones ; metabolism ; Humans ; K562 Cells ; Leukemia ; genetics ; metabolism ; pathology ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Triglycerides ; pharmacology