Objective: To investigate the effects of 5α - dihydrostestosterone (DHT) on calcium mobilization and growth of prostate cancer cell line LNCaP. Methods: Intracellular calcium concentration ([Ca2+]i) was assayed by MiraCal Image System using Fura-2/AM as Ca2+ fluorescence probe. Cell viability was observed by MTT assay and apoptosis by flow cytometry. Results: The calcium levels rapidly increased following addition of DHT, with the latency of response only in seconds. DHT at the concentrations of 1, 10, 100 and 1 000 nmol/L increased [Ca2+]i from (28±5), (29±5), (28±4) and (28±9) nmol/L to (31±3) (P>0.05,65±9) (P<0.01), (193±33) (P<0.001) and (208±42) nmol/L (P<0.001), respectively. The response induced by 1 000 nmol/L DHT was similar to that induced by 100 nmol/L DTH. DHT 1 000 nmol/L did not increase [Ca2+]i under extracellular Ca2+ -free condition. Blockers of L-type voltage-gated calcium channels, including verapamil (50 μmol/L), diltiazem (100 μmol/ L) or nifedipine (5 mmol/L) at 37°C for 5 min prior to stimulation with 1 000 nmol/L DHT, completely inhibited DHT-induced [Ca2+]i rise. Pre-treatment with inhibitor of phospholipase C such as neomycin sulfate (1 mmol/L) at 37°C for 3 min or inhibitor of ryanodine receptor such as procaine (50 mmol/L) at 37°C for 3 min had no influence on [Ca2+]i rise induced by 1 000 nmol/L DHT. The optical density (D) values and early apoptosis rates of the cells stimulated with 1 000 nmol/L DHT for 48 h were significantly different from those of cells pre-treated with verapamil prior to stimulation with 1 000 nmol/L DHT ([0.67±0.10]% vs [2.13±0.16] % and [14.31±2.29]% vs [1.07±0.19]%,P<0.01). Conclusion: DHT can induce rapid [Ca2+]i, rise in LNCaP cells in a concentration-dependent manner. The increase of [Ca2+]i, induced by DHT involves L-type voltage-gated calcium channels, but does not involve release of intracellular Ca2+ stores. The increase of [Ca2+]i induced by DHT increases apoptosis and inhibits growth of LNCaP cells.

Objective: To investigate the effect of human bone marrow mesenchymal stem cells (MSCs) conditioned medium (CM) on cell growth of prostate cancer cell line PC-3. Methods: MSCs were isolated and culture-expanded by using density gradient centrifugation from a healthy adult. When the passage 3 (P3) culture reached 80% confluence, the medium was changed to fresh medium. After 24-hour incubation, the supernatant (MSCs-CM) was collected. Then the PC-3 cells were incubated in fresh medium alone (as the control group) or in the medium supplemented with 50% MSCs-CM (as the CM group). The ultrastructure of PC-3 cells was observed under a transmission electron microscope. The cell viability was measured by WST-8 assay, and the cell cycle was analyzed by flow cytometry (FCM). Results: On day 4, the nucleus to cytoplasm ratio was lower and the number of organelles was much more in PC-3 cells in the CM group than those in the control group. The WST-8 assay showed that the absorbance values (on day 1, 3, 5 and 7) of the CM group were significantly higher than those in the control group (P<0.01), except for the value on day 1 (P>0.05). On day 4, the percentage of cells in G0/G1 phase was decreased while which in S+G2/M phase was increased in the CM group compared with those in the control group (56.20%±0.78% vs 68.54%±1.21%, P<0.01; 43.80%±0.77% vs 31.46%±1.20%, P<0.01). Conclusion: MSCs-CM can promote the proliferation of PC-3 cells through accelerating G 1 to S phase transition. Copyright© 2011 by TUMOR.

Objective To assess the volume of epicardial adipose tissue(EAT) and intrathoracic adipose tissue(IAT) and the correlation of EAT,IAT and the EAT/IAT ratio with the severity of coronary artery atherosclerosis in patients with metabolic syndrome(MS) and coronary heart disease(CHD).Methods Ninty-seven MS subjects without coronary atherosclerosis and one hundred and eighteen MS subjects with CHD were enrolled in this study.The volumes of EAT,PAT and IAT were measured using axial data from base to apex traced manually with a dedicated semiautomatic software program-volume.Results (1)Compared with MS subjects without coronary atherosclerosis,MS subjects with CHD had significantly increased age[(60.6± 1 1.4)years vs.(57.9 ± 8.7) years,P=0.001],positive family history of cardiovascular disease(29.7％ vs.21.6％,P=0.03),EAT[(98.3±41.4) cm3 vs.(82.2±39.7) cm3,P=0.001],IAT [(171.3±64.1) cm3 vs.(156.2±48.1) cm3,P=0.001] and HbA1c[(7.1 ± 1.8)％ vs.(6.6±2.3)％,P=0.02],but siginificantly reduced total cholesterol[TC,(4.9 ± 1.2)mmol/L vs.(5.4 ± 1.0) mmol/L,P=0.003],low-density lipoprotein cholesterol[LDL-C,(3.0±1.1) mmol/L vs.(3.6±1.0) mmol/L,P=0.03] and high-density lipoprotein cholesterol[HDL-C,(1.0 ± 0.4) mmol/L vs.(1.1 ± 0.3) mmol/L,P=0.04].(2) In MS subjects without coronary atherosclerosis,the volumes of EAT,pericardial adipose tissue(PAT) and IAT were associated with gender,body mass index(BMI),waist circumference(WC),diabetes mellitus and hyperlipidemia(all P＜0.05); however,in MS subjects with CHD,the volumes of EAT,PAT and IAT were associated with BMI and WC (both P＜0.05).(3) The volumes of EAT and IAT were correlated with calcification grades(r values were 0.45 and 0.50,P values were 0.017 and 0.013,respectively) and Gensini score(r values were 0.476 and 0.563,P values were 0.015 and 0.017,respectively) instead of coronary artery calcifacation score(CACS).Moreover,the EAT/IAT ratio was negatively correlated with CACS(r=-0.321,P=0.028).Conclusions Compared with MS subjects without coronary atherosclerosis,MS subjects with CHD have significantly increased EAT and IAT volumes.In our subjects,the volumes of EAT,PAT and IAT were associated with BMI and WC.Only in MS subjects with CHD,the volumes of EAT and IAT were correlated with caleification and Gensini score.

Objective To investigate the correlation between the angle and distribution of non-calcified plaque of left main coronary bifurcation and its hydrodynamic theory. Methods Retrospective analysis of 55 patients suspected of coronary heart disease who had undergone coronary computed tomographic angiography from January 2014 to June 2016.Angle,quadrant distribution,distance from bifurcation reference plane,minimal lumen area and extravascular elastic membrane cross section area of left main coronary bifurcation were measured. Then we calculated the plaque burden. The left main coronary bifurcation lesions were divided into plaque group and non-plaque group.Student's t test was used to compare the two groups.The left main coronary bifurcation lesions were divided into≥70°group and<70° group, Chi-square test and Fisher exact test were used to compared the two groups. Pearson correlation analysis was used to analyse the bifurcation angle and plaque distance. The data of non-plaque group was imported into the MIMICS,3-MATIC and abaqus CFD software to establish the hydrodynamic model,which was used to analyze the changes of pressure and velocity at different angles.Results Plaque group has 25 patients whose average value of the bifurcation angle was(55.95±14.77)°;non-plaque group has 30 patients whose average value of the bifurcation angle was(55.95±14.77)°.Angle difference was statistically significant between plaque and non-plaque group(t=2.893,P=0.006);Plaque difference was statistically significant between≥70°and<70°groups.The distance was negatively correlated with bifurcation angle(r=-0.773,P<0.001). Following the increase of bifurcation angle, the vascular wall area of high pressure and low speed region growed The green region of low-wall pressure with irregular distribution appeared in the lateral wall of the vessel that opposite to the red region of the bifurcation,and this region had lager gradient variability of wall pressure. Conclusion With the increase of coronary bifurcation angle, the possibility of plaque formation increases, the distance from plaque to bifurcation increases,the plaques tend to appear in the lateral wall of the blood vessels.

Developmental changes in children can affect drug disposition and clinical effects. A physiologically-based pharmacokinetic (PBPK) model is a mathematical model that can be used to predict blood drug concentrations in children and gain insight into age-dependent physiological differences in drug disposition impact. Pediatric PBPK (P-PBPK) models have attracted attention over the past decade. With the concerted efforts of academia, pharmaceutical companies, and regulatory agencies, there are more and more examples of pediatric clinical studies using PBPK models. Nevertheless, the number of P-PBPK models and their predictive performance still lag behind adult models. By referring to the literature, we study the process of children adapting to adult absorption, distribution, metabolism, and excretion (ADME) parameters and analyze the general principles of P-PBPK model establishment. In addition, we summarize the functions and application examples of commonly used P-PBPK modeling software to provide a basis for the rational application of modeling software.

OBJECTIVETo explore the efficacy differences in early intervention of different acupuncture and moxibustion methods on gastrocnemius fatigue in rats induced by electrical stimulation.

METHODSFifty male SD rats were randomly divided into a control group, a model group, a hand acupuncture group, an electroacupuncture group and a moxibustion group, 10 rats in each group. Electrical stimulation of the sciatic nerve was given in the control group and gastrocnemius fatigue test was induced by electrical stimulation of the sciatic nerve in the model group after anesthesia without other treatment, but just take 6 times interval stimulation in the control group. The hand acupuncture group, the electroacupuncture group and the moxibustion group were treated with the corresponding acupoints stimulation method respectively for 20 min before gastrocnemius fatigue test, and Dazhui (GV 14) and Zusanli (ST 36) were selected. Immediately after gastrocnemius fatigue test, three or four gastrocnemius tissues at the same site on the right side were quickly taken for making specimen for transmission electron microscope (TEM). The changes of skeletal muscle ultrastructure of myofibrils, mitochondria, sarcoplasmic reticulum, glycogen particles were observed under TEM.

RESULTS(1) Muscle fibers disorder, partial mitochondrial vacuolization and glycogen particles smaller were shown in the model group. (2) No abnormalities were shown in the hand acupuncture group and the moxibustion group with mitochondrial morphology and number, which better than that in the model group, and glycogen particles increased. (3) Abnormal changes in morphology were shown in the electroacupuncture group with part of the muscle fibers derangement, Z line malalignment and a few mitochondria vacuolization.

CONCLUSIONHand acupuncture, electroacupuncture and moxibustion have the different effects on ultrastructure of gastrocoemius in rats. Acupuncture and moxibustion have shown good effects on the prevention and treatment of exercise-induced skeletal muscle cell and organelle damage and delaying exercise-induced fatigue.

Acupuncture Points ; Acupuncture Therapy ; Animals ; Glycogen ; metabolism ; Male ; Mitochondria ; metabolism ; ultrastructure ; Moxibustion ; Muscle, Skeletal ; metabolism ; ultrastructure ; Rats ; Rats, Sprague-Dawley

Objective To investigate in vitro porcine myocardial shear modulus using real-time shear wave elastography.Methods Shear wave elastography was used on four in vitro porcine hearts.The probe was placed parallel to the long or short-axis of the heart on the anterior wall of the left ventricle,and myocardial shear modulus were measured in subepicardial,middle,subendocardial layers,respectively.Results Shear modulus of subepicardial,middle,subendocardial myocardium were (46.04 ± 17.07)kPa,(87.70 ± 29.67) kPa,( 115.73 ± 30.04) kPa,respectively,when the probe was placed parallel to the long-axis of the heart; whereas those were (78.71 ± 26.48) kPa,(77.08 ± 34.00)kPa,(70.69 ± 41.38) kPa,respectively,when the probe was placed parallel to the short-axis of the heart.Conclusions By using realtime shear wave elastography,the shear modulus values measured in subepicardial,middle,subendocardial myocardium of the left ventricle are different,even myocardium in the same location appears different shear modulus values when the probe orientation are different.

To develop a core-shell structure pDNA-CaPi-PLGA nanoparticles (CS-pDNA-CaPi-PLGA-NPs), calcium phosphate-pDNA nano complexes (CaPi-pDNA) were encapsulated inside of PLGA shells. The characteristics of the nanoparticles, including morphology, average particle size, zeta potential, entrapment efficiency, loading efficiency, stability in medium, pDNA protection ability from nuclease degradation, in vitro release, cytotoxicity and cell transfection were investigated and compared with the embedded structured CaPi modified PLGA nanoparticles (embedded-pDNA-CaPi-PLGA-NPs). The results showed that the obtained CS-pDNA-CaPi-PLGA-NPs were spherical in shape with an average particle size of (155 +/- 4.5) nm, zeta potentials of (-0.38 +/- 0.1) mV, entrapment efficiency of (80.56 +/- 2.5)% and loading efficiency of (1.16 +/- 0.04)%. The CS-pDNA-CaPi-PLGA-NPs were stable in the release media and could protect pDNA against nuclease degradation. And they also exhibited sustained release of pDNA in vitro. The highest gene transfection efficiency of the CS-pDNA-CaPi-PLGA-NPs in vitro reached (24.66 +/- 0.46)% (after 72 h transfection), which was significantly higher than that of free pDNA [(0.33 +/- 0.04)%, P < 0.01] and the pDNA-PLGA-NPs [(1.5 +/- 0.07)%, P < 0.01]. Besides, the transfection lasted for longer time than that of embedded-pDNA-CaPi-PLGA-NPs and the cytotoxicity of it was significantly lower than that of PEI (P < 0.01). These results indicate that CS-pDNA-CaPi-PLGA-NPs are a promising non-viral gene vector. Key words: gene delivery system; polylactic-co-glycolic acid; calcium phosphate; nanoparticle

Objective To establish the cell model of intractable epilepsy and to observe its neuronal damage and morphologic change of neurites.Methods The model was established by exposing hippocampal neurons to Mg2+ -free media for 3 hours on days 10 of culture.Expression of lactic acid dehydrogenase (LDH) in supernatant was measured as an index of neuronal damage.The morphologic change of neurons and neurites was observed by optical microscope and scanning electron microscope (SEM).Results Compared to the control group, level of LDH (U/L) was significantly increased in the model group at different time points (3 hours: 4.26 ± 1.28, 6 hours: 6.56 ±2.34 and 24 hours: 16.67 ±3.57, P <0.05).With time prolonging, release of LDH in the model group was gradually increased (F = 39.316,P <0.05).Under optical microscope, neurons of model group migrated closely to each other and neurite connections appeared to be gradually "reticulated" after Mg2+ -free media treatment for 24 hours; and the "reticulated" neurites connections become more obvious after 72 hours.Under SEM, neuronal membrane was rough and had several small depressions, neurites were interlaced in cluster.Conclusions Neuronal damage and morphologic change of neurites are verified in the cell model of intractable epilepsy.

ObjectiveTo analyze the differential expression of microRNA (miRNA) and its significance in patients with neonatal necrotizing enterocolitis (NEC).MethodsTwenty-five patients diagnosed with NEC with Bell stage≥Ⅱ, and 25 non-NEC patients as control group admitted to Guangzhou Women and Children's Medical Center between October 2014 and November 2015 were collected. White blood cells were extracted from the peripheral blood. Five samples were selected randomly each from NEC group and control group, and sequenced by second-generation Illumina high-throughput sequencing, screened for differentially expressed miRNA and analyzed for target genes prediction and biological function. The rest samples of the two groups were detected by real-time fluorescent quantitative polymerase chain reaction technology (RT-qPCR), the results were used to validate the results of high-throughput sequencing. Differentially expressed miRNA in the two groups of data was analyzed using DEGseq software.P<0.05 was considered statistically significant.P<0.01,q<0.001 and丨Log2 Ratio丨≥1 were taken as criteria for screening the differential expression. The differential expressions of miRNA in NEC group and control group were analyzed by cluster analysis using MeV4.6 software.ResultsA total of 482 miRNAs were differentially expressed in the two groups, with significant difference (P<0.05). Among them, 126 were known miRNAs with significantly differential expression in the two groups, with 58 being up-regulated and 68 being down-regulated. The results of up-regulated miRNAs (hsa-miR-223-5p,-183-3p,-222-5p) and down-regulated miRNAs (hsa-miR-23b-5p,-150-5p,-146a-3p,-1298-5p) were confirmed to be consistent with the results of sequencing. Bioinformatics analysis showed that the target genes with differential miRNA expression mainly involved Toll-like receptor signal transduction pathway, mitogen-activated protein kinase pathway, JAK-STAT and other signal transduction pathways.ConclusionsThere are significantly differential expressions of miRNAs in peripheral white blood cells of NEC neonates. These miRNs may be involved in the occurrence and development of NEC via adjusting different target genes to regulate the signal pathway.