Child ; Collagen Type IV ; Female ; Humans ; Immunohistochemistry ; methods ; Kidney ; pathology ; Male ; Nephritis, Hereditary ; diagnosis ; pathology

OBJECTIVETo investigate phase I and phase II enzyme activities in drug metabolism with tissue-like cultures of rat hepatocytes.

METHODSThe gel entrapment and spheroid culture of hepatocytes were used as tissue-like cultures and the monolayer culture was used as a control. The metabolism of phenacetin and 7-hydroxycoumarin (7-HC) was evaluated as the activities of phase I and phase II enzymes after incubated in medium for a period of time. The metabolites were assayed by HPLC. The hepatocytes were exposed to beta-naphthoflavone (BNF, 50 micromol x L(-1)) before the phase I and phase II enzyme activities were analyzed.

RESULTIn monolayer culture, phase I parameters decreased quickly and did not detected at d 5, and the phase II enzyme activities were not detected at d 7. In other two models of tissue-like cultures, the activities of phase I and phase II enzyme maintained at 32%-50% of the initial value at d 7. Paracetamol formation rates in spheroid culture maintained at 96% of that at d 1. The phase I enzyme activities of the spheroid culture were maintained from d 1 to d 3 at a level of 2.7-3.9-fold higher than the monolayer culture. After exposure to BNF the activities on phase I enzyme increased by about 2.5-fold (P <0.05) in all three culture models, while the increase in phase II enzyme was not significant.

CONCLUSIONThe gel entrapment culture and spheroid culture are superior to the monolayer culture in maintenance of drug metabolic enzyme activities.

Animals ; Cells, Cultured ; Cytochrome P-450 Enzyme System ; metabolism ; Enzyme Activation ; Female ; Hepatocytes ; cytology ; Male ; Phenacetin ; metabolism ; Rats ; Rats, Sprague-Dawley ; Umbelliferones ; metabolism ; beta-Naphthoflavone ; pharmacology

Nitrites play multiple characteristic functions in invasion and metastasis of hepatic cancer cells, but the exact mechanism is not yet known. Cancer cells can maintain the malignant characteristics via clearance of excess mitochondria by mitophagy. The purpose of this article was to determine the roles of nitrite, reactive oxygen species (ROS) and hypoxia inducing factor 1 alpha (HIF-1 α) in mitophagy of hepatic cancer cells. After exposure of human hepatocellular carcinoma SMMC-7721 cells to a serial concentrations of sodium nitrite for 24 h under normal oxygen, the maximal cell vitality was increased by 16 mg x (-1) sodium nitrite. In addition, the potentials of migration and invasion for SMMC-7721 cells were increased significantly at the same time. Furthermore, sodium nitrite exposure displayed an increase of stress fibers, lamellipodum and perinuclear mitochondrial distribution by cell staining with Actin-Tracker Green and Mito-Tracker Red, which was reversed by N-acetylcysteine (NAC, a reactive oxygen scavenger). DCFH-DA staining with fluorescent microscopy showed that the intracellular level of ROS concentration was increased by the sodium nitrite treatment. LC3 immunostaining and Western blot results showed that sodium nitrite enhanced cell autophagy flux. Under the transmission electron microscopy (TEM), more autolysosomes formed after sodium nitrite treatment and NAC could prevent autophagosome degradation. RT-PCR results indicated that the expression levels of COX I and COXIV mRNA were decreased significantly after sodium nitrite treatment. Meanwhile, laser scanning confocal microscopy showed that sodium nitrite significantly reduced mitochondrial mass detected by Mito-Tracker Green staining. The expression levels of HIF-1α, Beclin-1 and Bnip3 (mitophagy marker molecular) increased remarkably after sodium nitrite treatment, which were reversed by NAC. Our results demonstrated that sodium nitrite (16 mg x L(-1)) increased the potentials of invasion and migration of hepatic cancer SMMC-7721 cells through induction of ROS and HIF-1α mediated mitophagy.
Acetylcysteine ; pharmacology ; Autophagy ; Carcinoma, Hepatocellular ; pathology ; Cell Line, Tumor ; Cell Movement ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Liver Neoplasms ; pathology ; Mitochondrial Degradation ; Neoplasm Invasiveness ; Nitrites ; metabolism ; Reactive Oxygen Species ; metabolism ; Sodium Nitrite ; pharmacology

Six kinds of saponins (I, II, VII, PA, H) content of 22 samples of Paris polyphylla var. yunnanensis collected from different regions of Yunnan province were determined by HPLC, data was analyzed by SPSS 17. The results showed that the effect of altitude on saponin content was not significant, and the effect of growth area of saponins in P. polyphylla var. yunnanensis was significant, saponin content in sample from west Yunnan was significantly higher than that of samples from other regions.
Altitude ; China ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; analysis ; Liliaceae ; chemistry ; Saponins ; analysis

Objective To prospectively compare MR pulmonary perfusion imaging with quantitative HRCT for the detection of mild chronic obstructive pulmonary disease (COPD) and classification of COPD.Methods Sixty-two consecutive patients with COPD and 17 healthy volunteers underwent pulmonary function test (PFT),HRCT and MR perfusion imaging on the same day.According to the Global Initiative for Chronic Obstructive Lung Disease (GOLD),all COPD patients were classified into 4 stages:stage Ⅰ (n =19),stage Ⅱ (n =17),stage Ⅲ (n =14),stage Ⅳ (n =12).The signal intensity of perfusion defects (SIPD),signal intensity of normal lung perfusion (SInormal) on 3D MR perfusion were obtained through postprocessing and the signal intensity ratio (RSI) was calculated.The total lung volume (TLV) was calculated automatically on HRCT and the total emphysema volume (TEV) was obtained by applying -950 HU thresholds.The TEV/TLV was deduced as emphysema index (EI).Several comparisons were made between the volunteers and COPD patients by one-way ANOVA and Kruskal-Wallis test.Results The RSI,SIPD,PEI,MSI,MSD values of MRI perfusion in volunteers (43.9 ± 7.2,48.2 ± 19.7,31.4,55.7,44.1) were significantly different from those in patients with COPD (18.1 ± 8.1,47.4 ± 20.0,8.6,30.2,22.7) (P ＜ 0.01).The RSI showed a significant difference between stage Ⅰ (24.4 ± 9.8) and stage Ⅲ (15.9 ± 5.3) or Ⅳ (9.2 ± 2.7) and between stage Ⅱ (19.9 ± 3.1) and stage Ⅳ (t =4.05-6.64,P ＜0.01).However,all MRI perfusion parameters between stage Ⅰ and stage Ⅱ,stage Ⅱ and stage Ⅲ,stage Ⅲ and stage Ⅳ were no differences (t =2.00-4.46,P ＞ 0.05).The median of EI in volunteers and stage Ⅰ-Ⅳ COPD patients were 1.2,3.8,8.0,13.7,18.3,and the quartile range were 3.7,7.1,9.2,10.5,7.7,respectively.The EI in volunteers showed significant differences with that of stage Ⅱ-Ⅳ COPD and the EI of stage Ⅳ was different from that of stage Ⅱ or Ⅰ (t =-7.32--1.85,P ＜ 0.01),but there was no significant difference between volunteers and stage Ⅰ COPD (t =-1.99,P ＞ 0.05).Conclusions The RSI of MRI is more sensitive than that of HRCT for assessing mild COPD.The severity of COPD could be reflected by MRI perfusion and HRCT.

The authors helpfully discuss the design idea,experimental module design,examination methods,and experiment textbook construction in experimental teaching of microbiology,and conduct further researches on the basic skill training,verifying experiment,integrative experiment,and investigative experiment in the course. This study aims to enhance effects of the experimental teaching,to cultivate high potential talents who can master essential knowledge and skills,and creatively carry out scientific research.

Objective To evaluate the histologic changes in the dermis and the changes of the rate of type Ⅲ and type Ⅰ collagen by the radiofrequency device. Methods The effects of radiofrequency current on the dermis were observed. Ten rabbits were treated by radiofrequency, and the histologic change in the dermis were observed by H-E staining and Sirius red staining. Results After RF treatment, the fibers in the dermis appeared more compact and the quantity of the type Ⅲ (red) and type Ⅰ (green) collagen were both increased. The fibers in the dermis appeared more compact and the rate of type Ⅲ and type Ⅰ collagen was increased. It was also found that a significant proliferation of dermal collagen was observed in 8 days after treatment. As time went by, the proliferation of dermal collagen was more pronounced, and the rate of type Ⅲ was increased. Conclusion The radiofrequency current can increase the quantity of collagen in the dermis and increase the rate of type Ⅲ and type Ⅰ collagen, which may be one of the key mechanisms of facial rejuvenation by RF.

Objective To investigate the diagnostic technique of Alport syndrome(AS)by immunohistochemical staining of type Ⅳ collagen ? chains on paraffin-embedded renal sections.Methods Renal biopsies were obtained from 2 patients with autosomal recessive form of AS,2 female patients and 2 male patients with X-linked dominant form of AS and 2 patients with hematuria(1male and 1 female).AS was diagnosed according to symptoms,family history,pathology,immunofluorescence staining of type Ⅳ collagen ? chains on renal and skin biopsies and gene analysis.Normal portions of nephrectomized kidneys from 2 patients with renal tumor were used as controls.Type Ⅳ collagen ? chains were stained by two-step immunohistochemistry staining method on paraffin-embedded renal sections.Three antigen retrieval methods including autoclave heating,pepsin digestion and proteinase were investigated to find the best antigen retrieval method for type Ⅳ collagen ? chains.The findings were compared with those examined by immunofluorescence staining on fresh frozen sections.Results By immunohistochemistry staining,type Ⅳ collagen ?3 and ?5 chains showed continuous linear pattern along glomerular basement membrane on sections from the controls and the hematuria patients,intermittent linear pattern for X-linked dominant female AS patients,negative for X-linked dominant male AS patient.For patients with autosomal recessive AS,the staining of type Ⅳ collagen ?3 and ?5 chains were negative on glomerular basement membrane,but ?5 chain was positive on glomerular capsules and partial tubular basement membrane.The results were the same as those examined by immunofluorescence staining.Conclusion AS can be diagnosed by immunohistochemistry staining of type Ⅳ collagen on paraffin-embedded renal sections,which is a new technique for diagnosis of AS in China.

Objective To identify the polyreactivity of a monoclonal natural anti-keratin autoantibody 3B4 and to analyze its possible molecular me chanism.Methods enzyme-linked immunosorbent assay (ELISA)and immunohistoche mistry were applied to test the binding reactivity of 3B4 against different anti gens and tissues.The variable region genes and their amino acid composition wer e sequenced.Results 3B4 could reacted with a range of antigens and tissues,i n addition to keratin and skin.The variable region genes of its light chain and heavy chain showed high homology with germline genes VK1 am4 and VH1 J558.42.H CDR3 region,which mainly composed of short side chain amino acids(from 294 to 324 nucleotides around the heavy chain),was the only motif that differs from ot her highly homologous immunoglobulin genes.Conclusions The monoclonal natural anti-keratin autoantibody 3B4,with its variable region genes highly homologo us to germline genes,is highly polyreactive.The flexibility of HCDR3 may contr ibute to the polyreactivity.

This study investigated the effect of arctigenin (Arc) on the cell activation, cytokines expression, proliferation, and cell-cycle distribution of mouse T lymphocytes. Mouse lymphocytes were prepared from lymph node and treated with Phorbol-12-myristate-13-acetate (PMA)/Ionimycin (Ion) and/or Arc. CD69, CD25, cytokines, proliferation and cell cycle were assayed by flow cytometry. The results showed that, at concentrations of less than 1.00 micromol x L(-1), Arc expressed non-obvious cell damage to cultured lymphocytes, however, it could significantly down-regulate the expression of CD69 and CD25, as well as TNF-alpha, IFN-gamma, IL-2, IL-4, IL-6 and IL-10 on PMA/Ion stimulated lymphocytes. At the same time, Arc could also inhibit the proliferation of PMA/Ion-activated lymphocytes and exhibited lymphocyte G 0/G1 phase cycle arrest. These results suggest that Arc possesses significant anti-inflammatory effects that may be mediated through the regulation of cell activation, cytokines expression and cell proliferation.
Animals ; Anti-Inflammatory Agents ; isolation & purification ; pharmacology ; Antigens, CD ; metabolism ; Antigens, Differentiation, T-Lymphocyte ; metabolism ; Arctium ; chemistry ; Cell Cycle Checkpoints ; drug effects ; Cell Proliferation ; drug effects ; Cytokines ; metabolism ; Female ; Furans ; isolation & purification ; pharmacology ; Interferon-gamma ; metabolism ; Interleukin-10 ; metabolism ; Interleukin-2 ; metabolism ; Interleukin-2 Receptor alpha Subunit ; metabolism ; Interleukin-4 ; metabolism ; Interleukin-6 ; metabolism ; Ionomycin ; pharmacology ; Lectins, C-Type ; metabolism ; Lignans ; isolation & purification ; pharmacology ; Lymphocyte Activation ; drug effects ; Mice ; Mice, Inbred BALB C ; Plants, Medicinal ; chemistry ; T-Lymphocytes ; cytology ; drug effects ; immunology ; Tetradecanoylphorbol Acetate ; pharmacology ; Tumor Necrosis Factor-alpha ; metabolism