Hydrolytic amino acids were extracted by acid hydrolysis method, then derivatized with phenyl isothiocyanate (PITC). And the samples were analysed by HPLC on an Ultimate Prime C18 (4.6 mm x 250 mm, 5 microm) column with gradient elution of 0.1 mol x L(-1) sodium acetate buffer solution (adjusted to pH 6. 5)-acetonitrile (93:7) (A) and acetonitrile-water (8:2) (B) at a flow rate of 1.0 mL x min(-1). Column temperature was 40 degrees C and the detected wavelength was 254 nm. Amino acids derivative solution remained stable in 36 hours. The response was linear for 16 amino acids with a correlation coefficient r > 0.999 5. The average recoveries were 98.01% -101.8%. The method is reliable with good accuracy and repeatability, which is useful for the determination of amino acids in Galli Gigerii Endothelium Corneum.
Amino Acids ; analysis ; Animals ; Chickens ; Chromatography, High Pressure Liquid ; Chromatography, Reverse-Phase ; Endothelium ; chemistry ; Gizzard, Avian ; chemistry

OBJECTIVETo analyze the relationship between the pulse pressure (PP) and atherosclerotic renal artery stenosis (ARAS) in middle-age and eldery paients with hyperensio, so as to provide the basis for the diagnosis and treatment of ARAS.

METHODSWe tetro spectively analyzed the data of 257 patients with hypertension undergoing renal arteriography at General Hospital of PLA between 2009 and 2014. Their mean age was (64.09 +/- 7.81) years. According the results of arteriography, they were divided into two groups: ARAS group (n = 145), and non-ARAS group (n = 112). Their PP and multinomial clinical indexes with ARAS were statistically analyzed.

RESULTSSingle factor correlation and Logisitic analysis showed that age, PP, serum creatinine (SCr) levels, and new onset of hypertension after 50 years of age were strongly positively related to ARAS. The age and PP and SCr was a team of continuous variable.

CONCLUSIONPP was the first new risk factor of ARAS in middile-aged and elderly patients with hypertension. It is suggested that PP may be a early predictive indicator and a new therapeutic target for ARAS.

Angiography ; Arteriosclerosis ; physiopathology ; Blood Pressure ; Humans ; Hypertension ; Middle Aged ; Renal Artery ; physiopathology ; Renal Artery Obstruction ; Risk Factors

OBJECTIVETo investigate the protective effect of overexpressed heme oxygenase-1 (HO-1) in hypoxia and the correlation between HO-1 overexpressoin and hypoxia inducible factor-1alpha (HIF-1alpha) in human hepatoma HepG2 cells.

METHODSThe expressions of HO-1 and HIF-1alpha mRNA as well as the protein were detected by RT-PCR and Western blotting, respectively. MTT assay was used to examine the relative cell survival rate. The total superoxide dismutase (SOD) activity was examined with a SOD kit.

RESULTSHypoxia induced overexpression of HO-1 gene in HepG2 cells at transcriptional and translational levels. The relative survival rate of HepG2 cells under hypoxia was significantly decreased after the HO-1 protein overexpression was inhibited by ZnPPIX (P < 0.01). The total SOD activity of cells was significantly increased after cells were treated by hypoxia for 16 hours (P < 0.05), while decreased significantly by HO-1 inhibitor ZnPPIX treatment (P < 0.01). HIF-1alpha was upregulated under hypoxia. In addition, the HO-1 overexpression under hypoxia was decreased by HIF-1alpha inhibitor, while the HIF-1alpha expression level under hypoxia was not significantly changed after HO-1 expression was inhibited by ZnPPIX.

CONCLUSIONThe overexpression of HO-1 in hypoxic HepG2 cells is HIF-1alpha-dependent or at least partly HIF-1alpha-dependent. The relative survival rate of hypoxic hepatoma cells was significantly decreased by HO-1 inhibitor treatment. The results of this study may offer new thought and drug target for the therapy of human hepatoma in the future.

Cell Hypoxia ; Cell Survival ; Heme Oxygenase-1 ; antagonists & inhibitors ; metabolism ; Hep G2 Cells ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; genetics ; metabolism ; Protoporphyrins ; pharmacology ; RNA, Messenger ; metabolism ; Superoxide Dismutase ; metabolism

OBJECTIVETo study and compare the effect of neutrophil elastase inhibitors (GW311616A and sivelestat) on the proliferation and apoptosis of U937 cells.

METHODSInhibitory effects of GW311616A and sivelestat on the proliferation of U937 cells were assayed by MTT assay. The morphologic changes of U937 cells were detected by transmission electron microscope, and apoptosis was observed by AnnexinV-FITC/PI staining. The changes of cell cycle and apoptosis were detected by flow cytometry. The expression of NE in U937 cells was observed by indirect immunofluorescence, the variations of content and activity of NE in U937 cells were measured through ELISA assay and colorimetric method.

RESULTSMTT showed that both NE inhibitors could inhibit the proliferation of U937 cells in a dose dependent manner. The IC50 of GW311616A and sivelestat were 150 and 214 μmol/L respectively. The inhibition effect of GW311616A was significantly higher than of sivelestat (P<0.01). Typical apoptosis morphological changes of U937 cells was observed through electron microscope. AnnexinV-FITC/PI staining showed that U937 cells could be induced to undergo apoptosis by the two inhibitors, the apoptosis ratio of 150μmol/L GW311616A group (13.60%) was significantly higher than that of 150μmol/L sivelestat group (3.69%)(P<0.01). The result of flow cytometry indicated that the apoptosis ratio of 150 μmol/L GW311616A group was 14.61%, U937 cell cycle was mainly blocked in G2/M phase; meanwhile 150 μmol/L sivelestat group as 4.25% with cell cycle in S phase. The fluorescence intensity of GW311616A group obviously decreased than of sivelestat group. And the two inhibitors could reduce the content and activity of NE in U937 cells, but the effect of GW311616A was significantly higher than of sivelestat (P<0.01).

CONCLUSIONGW311616A and sivelestat could inhibit the proliferation and cause apoptosis of U937 cells. Furthermore, GW311616A was more effective and harmful to cells than sivelestat.

Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Glycine ; analogs & derivatives ; pharmacology ; Humans ; Leukocyte Elastase ; antagonists & inhibitors ; Piperidines ; pharmacology ; Proteinase Inhibitory Proteins, Secretory ; pharmacology ; Sulfonamides ; pharmacology ; U937 Cells

Adenosine Triphosphatases ; genetics ; Adult ; Cation Transport Proteins ; genetics ; Copper-transporting ATPases ; Genotype ; Hepatolenticular Degeneration ; genetics ; Humans ; Liver ; metabolism ; Male ; RNA, Messenger ; analysis

Action Potentials ; Animals ; Electromyography ; Environmental Exposure ; Hypothermia ; Rats ; Rats, Wistar ; Seawater ; Thigh ; physiology ; physiopathology

OBJECTIVETo study the applied anatomy of the vascular and muscular innervations related to vascularized fibular grafts.

METHODSThirty-four cadaveric lower extremities were dissected for this study. The observations included fibular length, fibular nutrient artery, arcuate arteries, and innervation of fibular muscles. The fibulas were averagely divided into four segments and the locations of relevant vessels and nerves were ascertained.

RESULTSAll specimens had 1 fibular nutrient artery and 4-9 arcuate arteries except 1 specimen which had only 1 arcuate artery. The fibular nutrient artery and the first three arcuate arteries were constantly located between the distal half of the 1/4 segment and 2/4 segment of the fibula. The muscular branch of the superficial peroneal nerve passed through the surface of the periosteum in the 2/4 segment of the fibula.

CONCLUSIONSThe most proximal osteotomy point locates at the midpoint of the 1/4 segment by which it ensure the maximal potential for preserving the nutrient vessels. The muscular branch of the superficial peroneal nerve is fragile to injury at the 2/4 segment of the fibula.

Cadaver ; Female ; Fibula ; blood supply ; innervation ; Humans ; Male

Objective To investigate the etiology of patients with severe to profound hearing loss and to identify the ratio of hereditary hearing loss in Chffeng area in Northern China. Methods DNA were extracted from peripheral blood of 134 deaf patients from Chifeng special educational school and 100 normal hearing controls in Northern China. Audiology examinations showed that all patients had severe to profound bilateral sensorineural hearing impairment. Sequence analysis of the whole coding areas of GJB2, GJB3, GJB6, SLC26A4, mtDNA12SrRNA and mtDNAtRNASer(LCN) were performed. Individuals carrying SLC26A4 mutation were given further temporal bone CT scan. Results The ratio of hearing loss related to genetic factors in this population was 60. 45% (81/134). About 33.58% (45/134) of the patients were given accurate genetic diagnosis. GJB2 mutations were responsible for approximately 17.16% of the cases in ChiFeng area. By screening SLC26A4 followed by temporal bone CT scan, we diagnosed 20 cases of enlarged vestibular aqueduct (EVA) and/or other inner ear malformation. SLC26A4 mutations account for about 14. 93% of the cases. The aminoglycoside-related mtDNA 1555A > G mutation accounted for 0.76% of the eases in Chifeng area. In addition, another 13.43% (18/134)of the cases carried heterozygous GJB2 mutation and their hearing loss may be related to GJB2. 6. 72% (9/134)of the cases carried heterezygous SLC26A4 mutation who were not found EVA by temporal bone CT or not took CT examination for some reasons. However, their hearing loss may also be SLC26A4-related. About 2. 24% (3/134)of the cases carried mtDNA 12SrRNA 1095 T > C which may also be an aminoglycoside-related mutation and very likely be the cause of hearing loss. GJB3 might participate in the pathomechanism of hearing loss in 1.49% (2/134) of the patients. GJB6 mutation was not detected in this population. Conclusions The ratio of hearing loss related to genetic factors in the sample drawing population from Chifeng was 60. 45% (81cases). GJB2 is the most common gene and SLC26A4 is the second common gene next to GJB2 that cause deafness in this area.

BACKGROUNDIt is difficult and time-consuming for carrying out conventional hand-sewn bilioenteric anastomosis, especially for small bile duct anastomosis and laparoscopic procedure. In order to simplify it, we have developed a novel procedure of sutureless bilioenteric anastomosis with an intraluminal degradable stent. This study aimed to evaluate the feasibility and safety of this technique with cholangioduodenostomy in dog model.

METHODSA patent intraluminal degradable stent tube for sutureless choledochoduodenostomy in dog model was made with polylactic acid in diameter of 3 mm or 4 mm. Thirty-eight dogs were randomly divided into to a stent group (SG, n = 20) and a control group (CG, n = 18). Dogs in the SG underwent sutureless choledochoduodenostomy with intraluminal stent, while the CG underwent conventional choledochoduodenostomy (single layer discontinuous anastomosis with absorbable suture). Dogs of each group were divided into 4 subgroups according to time of death (1, 3, 6, and 12 months postoperatively) to evaluate the healing of anastomosis. Operation time, intraoperative tolerance pressure of anastomosis, rate of postoperative bile leakage, bursting pressure of anastomosis were compared between the two groups. Anastomosis tissue was observed afterwards by pathology evaluation, hydroxyproline content, serum bilirubin, liver enzyme level and magnetic resonance cholangio-pancreatography (MRCP) to assess the stricture.

RESULTSAll procedures were completed successfully. The surgical time of the SG was significantly less than the CG (SG: (19.2 ± 4.3) minutes, vs. CG: (29.2 ± 7.1) minutes, P = 0.000). One bile leakage was occurred in either group. No significant difference of intraoperative tolerance pressure of anastomosis, rate of bile leakage and postoperative bursting pressure of anastomosis, anastomotic stricture, hydroxyproline content, serum bilirubin and liver enzyme level was found between the two groups. MRCP showed no anastomosis stricture and obstruction during months of follow-up.

CONCLUSIONThe technique of sutureless choledochoduodenostomy with a degradable intraluminal stent is feasible and a safe procedure in this dog model.

Animals ; Choledochostomy ; methods ; Dogs ; Female ; Male ; Stents

BACKGROUNDTrichophyton rubrum represents the most common infectious fungus responsible for dermatophytosis in human, but the mechanism involved is still not completely understood. An appropriate model constructed to simulate host infection is the prerequisite to study the pathogenesis of dermatophytosis caused by T. rubrum. In this study, we intended to develop a new T. rubrum infection model in vitro, using the three-dimensional reconstructed epidermis - EpiSkin ®, and to pave the way for further investigation of the mechanisms involved in T. rubrum infection.

METHODSThe reconstructed human epidermis (RHE) was infected by inoculating low-dose (400 conidia) and high-dose (4000 conidia) T. rubrum conidia to optimize the infection dose. During the various periods after infection, the samples were processed for pathological examination and scanning electron microscopy (SEM) observation.

RESULTSThe histological analysis of RHE revealed a fully differentiated epidermis with a functional stratum corneum, which was analogous to the normal human epidermis. The results of hematoxylin and eosin staining and the periodic acid-Schiff staining showed that the infection dose of 400 conidia was in accord with the pathological characteristics of host dermatophytosis caused by T. rubrum. SEM observations further exhibited the process of T. rubrum infection in an intuitionistic way.

CONCLUSIONSWe established the T. rubrum infection model on RHE in vitro successfully. It is a promising model for further investigation of the mechanisms involved in T. rubrum infection.

Animals ; Disease Models, Animal ; Epidermis ; microbiology ; Humans ; Keratinocytes ; cytology ; Tissue Culture Techniques ; Trichophyton ; pathogenicity