Objective To develop a clinical laboratory vehicle with advantages in mobility,deployment,automation and informatization to meet the requirements of disaster relief.Methods Ford transit V348 ambulance was used as the base,which was equipped with routine blood analyzer,routine urine analyzer,biochemical analyzer and etc.The same instruments,agents and procedures were adopted as those in the clinical laboratory department of rear hospital,and LIS,air cleaning system and necessary auxiliary units were also involved in the modification.Results The clinical laboratory vehicle increased the working efficiency while decreased the errors.Conclusion The vehicle fulfills mobile medical support for military operations other than war such as disaster relief,and thus is worthy promoting practically.

AIMTo construct a plasmid carrying hepatocyte growth factor gene and investigate its effects in vitro.

METHODSA complementary DNA (cDNA) clone for human HGF was isolated from human placental cDNA, then subcloned into pUDK vector, which was constructed by ourselves, to form the pUDKH plasmid. The transfection efficiency and the expression level of HGF and VEGF were evaluated by transfecting pUDK or pUDKH into primary rat skeletal muscle cells. The biological effects of HGF-expressing product at different doses on endothelial cells were investigated in vitro, and assessed by MTT.

RESULTSThe primary rat skeletal muscle cells could be transfected efficiently with pUDKH (0.057%), and secreted HGF(16 -18 ng/4 x 10(5) cells) and VEGF proteins. The expressing product could significantly stimulate proliferation of human umbilical vein endothelial cells, in a dose-dependent manner (P < 0.05).

CONCLUSIONpUDKH has the potential application in vivo to treat ischemic diseases.

Animals ; Cells, Cultured ; DNA, Complementary ; genetics ; Genetic Vectors ; Hepatocyte Growth Factor ; genetics ; metabolism ; Humans ; Plasmids ; Rats ; Rats, Wistar ; Transfection

AIMTo investigate the protective effects of adenovirus-mediated hepatocyte growth factor (Ad-HGF) on injury of rat cortex neurons induced by in vitro serum-free culture.

METHODSFlow cytometry was used to assay the transfection rate of rat cortex neurons infected by adenovirus-mediated green fluorescent protein(Ad-GFP) at different multiplicity of infection (MOI) to find out the best MOI in experiment. ELISA was used to elucidate the expression patterns of cortex neuron. Neutral red stain and PI-Hoechst 33342 double stain were used to compare the viability of cortex neurons, which were cultured in serum-free medium for 6 h, 12 h, 24 h and 48 h respectively, among the Ad-HGF transfected group, the Ad-GFP transfected group and the control group.

RESULTSIt was found that when MOI was 50 PFU per cell, a transfection rate as high as 99.3% was maintained and Ad-HGF was able to express in cortex neurons effectively and persistently. In addition, the death rate and apoptotic rate of cortex neurons (infected 2 hours after seeding) cultured in serum-free medium for 12 h in Ad-HGF transfected group was significantly lower than that in both the Ad-GFP group and the control group (P < 0.05).

CONCLUSIONAd-HGF plays a protective role against in vitro serum-free culture induced injury on rat cortex neurons infected 2 hours after seeding. Though its effects on rat cortex neurons infected 5 days after seeding are not so remarkable, Ad-HGF also has the potential to protect cortex neurons from serum-free culture induced injury.

Adenoviridae ; genetics ; Animals ; Animals, Newborn ; Cells, Cultured ; Cerebral Cortex ; drug effects ; Culture Media, Serum-Free ; Hepatocyte Growth Factor ; genetics ; pharmacology ; Neurons ; drug effects ; Rats ; Rats, Wistar ; Transfection

Autophagy is an important metabolic mechanism involved in stress responses of cells such as inflamma -tion.A close molecular connection between autophagy and inflammation .On the other side , some inflammatory me-diators can regulate autophagy and its translation;Autophagy is able to suppress the excessive inflammation through various pathway .Therefore , deficient autophagy may associate with inflammatory diseases such as Crohn 's disease ( CD) .

BACKGROUNDHepatocyte growth factor (HGF) treats ischemic disease by promoting arteriogenesis, however, its mechanism of action is not known. The notch signaling pathway plays an important role in neovascularization. The relationship between the proliferation and migration ability of artery endothelial cells and the Dll4-Notch-Hey2 signaling pathway in the process of arteriogenesis was investigated as a mechanism of action of HGF.

METHODSBased on the prophase study cells and supernatant were harvested at the indicated time after human femoral artery endothelial cells (HFAECs) were infected with adenovirus-HGF (Ad-HGF) at 200 pfu/cell. Cells were analyzed for HGF expression and Notch1, Dll4 and Hey2 expression by ELISA and reverse transcription-PCR (RT-PCR). The changes in the proliferation and migration ability of HFAECs were observed by MTT and Transwell migration experiments. Ad-GFP-infected HFAECs were used as control.

RESULTSCompared with the control group the Ad-HGF group's HGF expression was not increased with time, and the induction by HGF of Notch1, Dll4 and Hey2 gene transcription was not enhanced with an increase of HGF. The proliferation ability of Ad-HGF-transduced HFAECs was enhanced and their migration ability was also enhanced in the presence of HGF.

CONCLUSIONSThrough activating the Dll4-Notch-Hey2 signaling pathway, HGF indirectly promotes the proliferation and migration ability of cells, so that offspring artery branches are formed.

Basic Helix-Loop-Helix Transcription Factors ; genetics ; metabolism ; Cell Line ; Enzyme-Linked Immunosorbent Assay ; Hepatocyte Growth Factor ; genetics ; metabolism ; Humans ; Intercellular Signaling Peptides and Proteins ; genetics ; metabolism ; Receptors, Notch ; genetics ; metabolism ; Repressor Proteins ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; genetics ; physiology

OBJECTIVETo review the effect of endothelial progenitor cells in neovascularization as well as their application to the therapy of tumors.

DATA SOURCESThe data used in this review were mainly from PubMed for relevant English language articles published from 1997 to 2009. The search term was "endothelial progenitor cells".

STUDY SELECTIONArticles regarding the role of endothelial progenitor cells in neovascularization and their application to the therapy of tumors were selected.

RESULTSEndothelial progenitor cells isolated from bone marrow, umbilical cord blood and peripheral blood can proliferate, mobilize and differentiate into mature endothelial cells. Experiments suggest endothelial progenitor cells take part in forming the tumor vascular through a variety of mechanisms related to vascular endothelial growth factor, matrix metalloproteinases, chemokine stromal cell-derived factor 1 and its receptor C-X-C receptor-4, erythropoietin, Notch signal pathway and so on. Evidence demonstrates that the number and function change of endothelial progenitor cells in peripheral blood can be used as a biomarker of the response of cancer patients to anti-tumor therapy and predict the prognosis and recurrence. In addition, irradiation temporarily increased endothelial cells number and decreased the endothelial progenitor cell counts in animal models. Meanwhile, in preclinical experiments, therapeutic gene-modified endothelial progenitor cells have been approved to attenuate tumor growth and offer a novel strategy for cell therapy and gene therapy of cancer.

CONCLUSIONSEndothelial progenitor cells play a particular role in neovascularization and have attractively potential prognostic and therapeutic applications to malignant tumors. However, a series of problems, such as the definitive biomarkers of endothelial progenitor cells, their interrelationship with radiotherapy and their application in cell therapy and gene therapy of tumors, need further investigation.

Animals ; Endothelial Cells ; physiology ; Extracellular Signal-Regulated MAP Kinases ; physiology ; Genetic Therapy ; Hematopoietic Stem Cell Mobilization ; Humans ; Neoplasms ; blood supply ; therapy ; Neovascularization, Pathologic ; etiology ; Neovascularization, Physiologic ; Receptors, CXCR4 ; physiology ; Stem Cells ; physiology

OBJECTIVETo carry out a genetic detection and analysis of Von Hippel-Lindau (VHL) gene in Chinese patients with sporadic pheochromocytoma.

METHODSDNA samples were extracted from peripheral blood cells and fresh pheochromocytoma specimens from 41 patients with sporadic pheochromocytoma were assayed by polymerase chain reaction and direct sequencing. The DNA samples of 50 healthy volunteers were extracted from peripheral blood as a control. The PCR products of exon 1, exon 2 and exon 3 were used for molecular analysis of the VHL gene. The genetic detection of family members of VHL gene mutations was also performed.

RESULTSOne of mutations was located at nucleotide 572 (G-->C) in exon 2, presenting a codon 120 from arginine (R) to threonine (T). Tow small insertions were locatated at nucleotide 623T (TTTGTtG) in exon 2, leading to a frameshift mutation. There were also three carriers of G572C and three carriers of 623T (TTTGTtG) in family members of the three cases.

CONCLUSIONThere are some Chinese patients with sporadic pheochromocytoma with tumorigenic VHL gene mutations. It is recommended to use the genetic detection and analysis of VHL gene as a routine examination for patients with sporadic pheoehromoeytoma under the age of 50 years with questionable family history. The genetic detection and analysis of VHL gene may be useful as a marker for the diagnosis of hereditary pheochromocytoma.

Adrenal Gland Neoplasms ; genetics ; Adult ; DNA Mutational Analysis ; DNA, Neoplasm ; genetics ; Exons ; Family ; Female ; Humans ; Male ; Middle Aged ; Mutation ; Pedigree ; Pheochromocytoma ; genetics ; Von Hippel-Lindau Tumor Suppressor Protein ; genetics

OBJECTIVETo explore the toxic effects of lead acetate on the apoptosis and ultrastructure of human renal tubular epithelial cells (HK-2).

METHODSAfter HK-2 cells were exposed to 5, 10 and 20 µmol/L lead acetate for 24 h, the morphological changes of HK-2 cells were observed by Hochest 33342-PI staining, and the ultrastructure changes of HK-2 cells were examined under a electron microscope, LDH activity and MDA content in supernatant of HK-2 cellular culture were detected by spectrophotometer, DNA damage of HK-2 was determined by DNA ladder and the apoptotic rates of HK-2 cells were measured by flow cytometry.

RESULTSThe morphological changes of apoptotic HK-2 cells in exposure group were observed by Hochest 33342-PI staining. The cytoplasm vacuoles, karyopycnosis, nuclear membrane vague and apoptotic bodies in HK-2 cells of exposure group were found under electron microscopy. LDH activity and MDA contents in exposure group increased significantly, as compared to control group (P < 0.01). The results of DNA Ladder showed that DNA damage of HK-2 cells in exposure group appeared. The apoptotic rates of HK-2 cells exposed to 5, 10, 20 µmol/L lead acetate were 14.16% ± 2.94%, 19.45% ± 2.73%, 25.01% ± 3.97%, respectively, which were significantly higher than that (5.81% ± 2.18%) in control group (P < 0.05).

CONCLUSIONLead acetate could remarkably induce the apoptosis of HK-2 cells and affect the kidney.

Apoptosis ; drug effects ; Cell Line ; Epithelial Cells ; cytology ; drug effects ; ultrastructure ; Humans ; Kidney Tubules, Proximal ; cytology ; drug effects ; ultrastructure ; Organometallic Compounds ; toxicity

OBJECTIVETo explore the effects of bone marrow-derived mesenchymal stem cells (BMSCs) transfected with adenoviral vector carrying hepatocyte growth factor (HGF, Ad-HGF) on burn wound healing.

METHODSBMSCs from male Wistar rats were separated and purified with Percoll separating medium by density gradient centrifugation and cultured with DMEM containing 20% fetal bovine serum (FBS). Then BMSCs were transfected with Ad-HGF at the optimal gene transduction efficiency of 100 multiplicity of infection (MOI). The efficiency of transfection and the expression of HGF in the suspension were detected by flow cytometry and enzyme linked immunosorbent assay (ELISA) respectively. Thirty-two female rats were subjected to 90 degree centigrade water for 12 seconds to induce a partial thickness skin burn. The animals were randomly divided into mesenchymal stem cells (MSCs) treatment group (Group A), Ad-HGF treatment group (Group B), Ad-HGF-modified MSCs treatment group (Group C) and saline control group (Group D). On days 3, 5, 7, 14 and 21 postburn, HE and Sirius red stain were performed to observe the burn wound healing and collagen content. The content of hydroxyproline in wounds was also detected. Transplanted cells and the expression of (sex-determining region Y) SRY gene were detected by in situ hybridization and polymerase chain reaction (PCR), while the expression of HGF in wound tissues was detected by ELISA.

RESULTSThe result of flow cytometry showed that the transfection efficiency was 86.41% at 100 MOI. Compared with the control group, the content of HGF in the supernatant after transfection increased time-dependently and peaked at 48 h, showing significant differences at 24 h, 48 h, 72 h and 96 h (P less than 0.01). Results of HE stain revealed that the range of re-epidermidalization in Group C was significantly larger than that in other groups in the first week. Three weeks postburn, the epidermis was significantly thicker in Group C than in other groups and the nails of dermis inserted into the derma of burn wounds. Sirius red stain showed that the content of collagen I in Group C was much less compared with that in other groups 21 days postburn. In situ hybridization revealed an expression of SRY gene in burned female rats, consistent with the finding of PCR. Immunohistochemistry demonstrated the largest increase of HGF expression in Group C, whose contents of hydroxyproline, however, decreased on day 7 postburn. Compared with other groups, the content of HGF in the wounds of Group C increased obviously on day 14 after transfection (P less than 0.05) and there was no significant difference among Groups A, B and D.

CONCLUSIONOur study suggests that transplantation of MSCs modified with Ad-HGF has positive effects on the healing of burn wounds probably through differentiation and release of relevant cytokines.

Adenoviridae ; genetics ; Animals ; Burns ; metabolism ; therapy ; Cells, Cultured ; Female ; Genetic Therapy ; Hepatocyte Growth Factor ; analysis ; genetics ; Humans ; Hydroxyproline ; analysis ; Male ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Rats ; Rats, Wistar ; Transfection ; Wound Healing

AIMTo assess whether hepatocyte growth factor recruits bone marrow-derived endothelial progenitor cells into blood circulation to participate in postnatal angiogenesis and endothelium repair.

METHODSThe adenovirus vector encoding HGF gene (Ad-HGF) were intravenous administrated into BALB/c mice, and then serum HGF was determined by enzyme-linked immunosorbent assay, the number of CD34+ cells in peripheral blood was assayed by flow cytometry, and the nucleated cells in peripheral blood were isolated, cultured and the endothelial cell colonies were characterized by staining with antibodies against tie-2, vWF. The carbon tetrachloride-induced liver damage model of female mice was established. The peripheral blood nucleated cells of Ad-HGF treated male mice were intravenous administrated into these mice, and 4 weeks later, in situ hybridization for the sry gene was used to identify the implanted cells in the damaged tissues.

RESULTSIntravenous administration of Ad-HGF resulted in significant elevation of serum hepatocyte growth factor level and induced profoundly increase of endothelial progenitor cells in the peripheral blood, which were characterized by their ability to form endothelial cell colonies in culture and expression of CD34, tie-2, and vW factor. HGF-mobilized endothelial progenitors could incorporate into sites of neovascularization in a liver regeneration model.

CONCLUSIONHepatocyte growth factor could markedly recruit bone marrow-derived endothelial progenitor cells into blood circulation.

Animals ; Bone Marrow Cells ; cytology ; Endothelial Cells ; cytology ; Female ; Hematopoietic Stem Cell Mobilization ; Hepatocyte Growth Factor ; pharmacology ; Male ; Mice ; Mice, Inbred BALB C ; Stem Cells ; cytology