Objective Taking methyl nonyl ketone as index to establish pharmacokinetic compartment model for study volatile oil from the whole herb of Houttuynia cordata by pharmacokinetic parameters in rabbits in vivo. Methods Using GC to determine the blood concentrations of methyl nonyl ketone in the volatile oil from whole herbs of H. cordata in rabbit plasma. The GC system used a capillary SPB-5 column and helium as carrier gas; the initial temperature was at 90 ℃, then raised to 190 ℃ by 10 ℃/min and kept for 5 min; the injector temperature was 250 ℃ and the FID detector temperature was 300 ℃; the injection volume was 5.0 ?L with splitless injection mode; n-hexane was used as solvent and n-pentadecane as interior standard substance. The blood concentration value of methyl nonyl ketone was fitted with 3P97 procedure. Results A capillary GC method was established for the detemination of methyl nonyl ketone in the volatile oil from whole herbs of H. cordata in rabbit plasma. Methyl nonyl ketone linearity was good (r=0.987 0) within 0.04—60.0 ?g/mL.The blood concentration-time course of precision in rabbit plasma conformed to two compartment model. Conclusion The experiment provides pharmacokinetic evidence for the rational administration and the further development of H. cordata and some methods for pharmacokinetic study of volatile oils from traditional Chinese medicinal materials.

Objective To explore the feasibility of using human umbilical cord derived mesenchymal stem cells as seed cells to repair sciatic nerve defects of rats by tissue engineering methods. Methods Mesenchymal stem cells from human umbilical cord were cultured and induced into neuron-liked cells,which were co-cultured with acellular basal lamina tube to construct tissue engineering nerve;models of sciatic nerve defects 10 mm in length were set up with thirty healthy adult SD rats and were divided randomly into 3 groups:tissue engineering nerve group (group A, compound of human umbilical cord derived mesenchymal stem cells and acellular basal lamina tube), pure acellular basal lamina tube group (group B), and autogenous nerve bridging group (group C). Evaluation of electrophysiological and histological results was carried out 10 weeks after operation. Results The engineering nerve group had good result in nerve regeneration which was close to the effect of autogenous nerve transfer group (group A), and much better than the effect of pure acellular basal lamina tube group. Conclusion Engineering nerves from human umbilical cord derived mesenchymal stem cells can effectively repair 10 mm defects of sciatic nerve.

Cell biology is a highly experimental discipline, and is a discipline with theoretical knowledge closely combined with practical operation.Most of the medical graduate students should master the cell biology theory in the graduate stage, and get on a series of cell biology researches.Therefore, it is important to master the most basic cell culture techniques.As a technical instructor in the cell culture laboratory, I have been summarizing and optimizing the teaching content and teaching mode during the past few years.The aim of the teaching work is to train the students' scientific attitude of seeking truth from facts so as to develop good experimental habits and to master the basic skills of cell culture.

OBJECTIVETo construct and characterize the TGF-β1, induced epithelial-mesenchymal transition (EMT) model of keloid epithelial cells in vitro, and to investigate the expression of epithelial stem cells related surface markers in keloid epithelial cells during EMT induction.

METHODSThe epithelial cells from 3 keloid samples of ears were cultured in vitro and induced by transforming growth factor betal (TGF-β1, 1 ng/ml) for 5 days, which was the experimental group, the same cells untreated were considered as the negative control group. The expressions of EMT-associated markers and regulative genes were detected using immunofluorescence staining, real-time PCR and western blot analysis. Then the surface markers of epithelial stem cells were detected using real-time PCR. Statistical significance was determined using Independent-Samples t Test, a p value less than 0. 05 was considered statistically significant.

RESULTSThe mRNA expression of transcription factor snail2 and mesenchymal-specific marker vimentin increased significantly in TGF-β1, induced keloid epithelial cells (P < 0. 05), in which snail2 increasing from 0. 91 ± 0. 23 to 1. 69 ± 0. 10, and vimentin from 5. 86 ± 2. 07 to 24. 29 ± 5. 39. Whereas the mRNA expression of epithelial-specific marker E-cadherin decreased from 1. 06 ± 0. 19 to 0. 65 ± 0. 09. The mRNA expression of CD29 and Lgr6, two surface markers of epithelial stem cells, significantly increased after induction of the TGF-β1, (P < 0. 05), from 0. 55 ± 0. 14 and 1. 61 ± 0. 31 to 1. 19 ± 0. 12 and 3. 84 t 0. 62 respectively. In induced cells, the immunofluorescence results showed staining of E- cadherin became faint, but the number of positive staining cells of vimentin increased. Western blot confirmed the protein expression of E-cadherin weakened, and the vimentin and p-Smad3 enhanced (P < 0. 05).

CONCLUSIONSTGF-β1, initiated EMT in keloid epithelial cells by inducing the up-regulation of snail2, and TGF-β1,/Smad3 signaling pathway was involved in EMT. EMT could change the phenotype of epithelial stem cells in keloid.

Biomarkers ; metabolism ; Cadherins ; genetics ; metabolism ; Epithelial Cells ; drug effects ; physiology ; Epithelial-Mesenchymal Transition ; drug effects ; physiology ; Humans ; In Vitro Techniques ; Keloid ; pathology ; RNA, Messenger ; metabolism ; Signal Transduction ; Smad3 Protein ; genetics ; metabolism ; Snail Family Transcription Factors ; Transcription Factors ; genetics ; metabolism ; Transforming Growth Factor beta1 ; metabolism ; pharmacology ; Up-Regulation ; Vimentin ; genetics ; metabolism

AIM: To explore the molecular mechanism of anti-shock effect of Shenmai (SM) injection and to observe its influence on TLR4 and I?B mRNA expression in hemorrhagic shock (HS) plus LPS rat lung. METHODS: Rats were divided into HS, HS+LPS, HS+LPS+ dexamethasone (DEX), HS+LPS+SM group, respectively. HS+LPS injury model of rats were induced by hemorrhagic shock (MAP 40 mmHg for 10 min) followed by intravenous administration of lipopolysaccharide (LPS, 1.5 mg/kg). Four hours later, the TNF-? contents in lung tissue were measured by ELISA, TLR4 mRNA and I?B? mRNA expression were assayed by RT-PCR, ultrastructural changes of the type Ⅱ alveolar cell were observed by electron microscope. RESULTS: The TLR4 mRNA expression level in HS+LPS+SM group was significantly decreased as compared to HS+LPS group (P

Objective To investigate the association of serum brain-derived neurotrophic factor (BDNF) and methylglyoxal (MG) levels with cognitive function in elderly patients with type 2 diabetes mellitus (T2DM). Methods The normal population and elderly patients with T2DM were frequency-matched by age, sex, and educational level. BDNF was detected by ELISA assay, MG by HPLC assay, and cognitive function by sets of repetitive mental state examination (RBANS) in the two groups. Results (1) Compared with control group, serum BDNF level in T2DM group was significantly decreased [ (4.97±3.05 vs 11.77±7.92)ng /ml, P<0.01]while serum MG level was elevated [(67.91 vs 43.86) nmol /L, P<0.05]. The increasing of serum MG was related to the decreasing of serum BDNF. (2) Compared with control group, the scores for standardized tests of cognitive scale, visual breadth, immediate memory, delayed memory, and attention areas in T2DM group were significantly decreased (all P<0.05). After the influencing factors were adjusted by multiple regression, the associations of serum BDNF level with cognitive scale standardized score, the delay associated with memory and attention functions were still evident, and serum MG level in T2DM group was still related with the levels of delayed memory, immediate memory, total scale standardization (all P<0.05). (3) Serum BDNF level was negatively correlated with serum MG level (P=0.031). Conclusions Cognitive function of elderly patients with T2DM is related with serum MG and BDNF levels. The increased serum MG as well as the decreasd serum BDNF levels maybe involved in the pathogenesis of impaired cognitive function.

Objective To investigate the role of human membrane associated sialidase (Neu3) in multidrug resistance ( MDR) of K562 cells. Methods K562 cells and K562/ADM cells were treated with DNR alone or with combination of DNR and NeuAC2en. The cell survival rate was measured by MTT assay; the expression of MDR related factors and apoptosis related factors were determined by Western blot and RT-PCR; Neu3 activity was detected by TBA reaction. Results The survival rate of K562/ADM cells was higher than that of K562 cells; NeuAC2en showed synergistic effect with DNR(P <0. 01) ; Neu3 activity of K562 and K562/ADM cells decreased after DNR or NeuAC2en induction and the decrease was most significant in the combination treatment group (P <0. 01). Under the identical condition, the protein expression of P-gp and Neu3 and the mRNA expression of MDR1, Neu3, BCL-xl and BCL-2 increased comparing with K562 cells. The mRNA level of BAX in K562 and K562/ADM groupsdid not changed significantly. MDR1, Neu3,BCL-xl and BCL-2 decreased after DNR induction and down-regulated most obviously in the groups treated by DNR combined with NeuAC2en. Conclusion Neu3 may be related with multidrug resistance of K562/ADM cell through regulating apoptosis and the expression of MDR1.

Objective To investigate the strategies of reducing the incidence of missed diagnosis of severe traumatic brain injuries combined with multiple trauma. Methods Data of 432 patients with severe traumatic brain injuries and multiple trauma (ISS≥20) from January 2000 to August 2007 were analyzed retrospectively. All patients were divided into missed diagnosis group (MD group, n =54) and non-missed diagnosis group (NMD group, n =378) for correlation analysis on ISS, GCS, anatomical locations of the missed diagnosis, the time of delayed diagnosis and the prognosis. Results ISS was (42.97±10.94) points in MD group, with statistical difference compared with NMD group (P < 0.05). The patients with GCS≤8 in MD group was more than those in NMD group (P < 0.05). Conclusions It is effective to prevent missed diagnosis and improve the survival of patients with severe traumatic brain injuries combined with multiple trauma by judging injury severity quickly and precisely based on the principle of "life first" and repeated and systemic physical examination.

AIM:To study the effect of vitamin K3(VK3) on the induction of apoptosis in androgen-independent prostate cancer cell PC-3M in vitro.METHODS:Cell viability was estimated by MTT assay.AO/EB staining was performed to detect apoptotic cells.Apoptosis and the changes of cell cycle were detected by flow cytometry.NAC was used to observe the effect of growth inhibition by VK3.RT-PCR was used to confirm the changes in gene expression.Levels of intracellular peroxides were estimated by using an oxidation-sensitive fluorescent probe DCFH-DA.RESULTS:PC-3M cells growth was significantly inhibited by VK3(≥60 ?mol/L,P

Objective:To investigate the effect of long-chain non coding RNA (lncrna) loc730101 in the proliferation, invasion and migration of U2OS cells, and its mechanism.Methods:From February, 2019 to October, 2019, U2OS cells cultured in vitro were divided into control group (normal culture), negative group (transfection nega- tive control), and interference group (transfection of interference sequences targeting LOC730101). The expression of LOC730101 in cells was detected by reverse transcription-polymerase chain reaction (RT-PCR). Cell proliferation ac tivity was tested by methylthiazolyldiphenyl-tetrazolium bromide (MTT) method. Cell clone formation rate was mearused by plate clone formation test. Cell cycle distribution was tested by flow cytometry. Cell invasion and migra- tion were examed by Transwell chamber. The expression levels of epithelial-mesenchymal transition-related proteins Vimentin, N-cadherin, E-cadherin, and Wnt/β-catenin signaling pathway related proteins in cells β-catenin, c-Myc, cyclin D1(CyclinD1) and matrix metalloproteinase-7(MMP-7) proteins were detected by Western blotting method. The date was statistical analysed and considered as statistically significant when P<0.05. Results:Compared with the control group, the expression level of LOC730101 (0.25±0.03 and 1.00±0.06) in interference group and control group, respectively. The same below), cell survival rate [(57.65±3.26)% and (100.00±7.39)%], clone formation rate [(13.03± 2.12)% and (25.35±3.58)%], number of invasive cells(51.36±3.48 and 92.85±6.62), number of migrating cells (77.15± 5.05 and 136.92±15.35), the percentage of cells in S phase [(20.54±2.15)% and (28.15±2.38)%] and G 2/M phase [(16.87±2.12)% and (23.36±3.12)%], as well as the expression level of Vimentin (0.52±0.04 and 1.17±0.13), N-cadherin (0.31±0.03 and 0.65±0.04), β-catenin (0.42±0.03 and 0.73±0.04), c-Myc (0.29±0.03 and 0.65±0.03), CyclinD1 (0.26± 0.02 and 0.58±0.04), MMP-7 protein (0.55±0.03 and 0.86±0.06) was decreased significantly ( P<0.05), while the per- centage of cells in G 0/G 1 phase [(62.62±5.15)% and (48.46±3.65)%] and the expression level of E-cadherin protein(0.82± 0.06 and 0.38±0.03) were increased significantly in the interference group ( P<0.05). But there was no significant differ- ence in each index in the negative group ( P>0.05). Conclusion:Reduce the regulation of LOC730101 can inhibit the proliferation, invasion and migration of U2OS cells, and its mechanism may be related to the inhibition of Wnt/β-catenin signaling pathway.