To explore the expression potential of heterogeneous genes using the backbone of infectious bronchitis virus (IBV) Beaudette strain, the ectodomain region of the Spike gene (1,302 bp) of IBV H120 strain was amplified by RT-PCR and replaced into the corresponding location of the IBV Beaudette strain full-length cDNA. This recombinant was designated as BeauR-H120(S1). BeauR-H120(S1) was directly used as the DNA template for the transcription of viral genomic RNA in vitro. Then, the transcription product was transfected into Vero cells by electroporation. At 48 h post-transfection, the transfected Vero cells were harvested, and passaging continued. A syncytium was not observed until the recombinant virus had passed through four passages. The presence of rBeau-H120(S1) was verified by the detection of the replaced ectodomain region of the H120 Spike gene using RT-PCR. Western blot analysis of rBeau-H120 (S1)-infected Vero cell lysates demonstrated that the nucleocapsid (N) protein was expressed, which implied that rBeau-H120(S1) could propagate in Vero cells. The TCIDs0 and EIDs0 data demonstrated that the titer levels of rBeau-H120(S1) reached 10(590+/-0.22)TCID50/mL and 10(6.13+/-0.23)EID50/mL in Vero cells and 9-day-old SPF chicken embryos, respectively. Protection studies showed that the percentage of antibody-positive chickens, which were vaccinated with rBeau-H120(S1) at 7 days after hatching, rose to 90% at 21 days post-inoculation. Inoculation provided an 85% rate of immune protection against a challenge of the virulent IBV M41 strain (103EID50/chicken). This recombinant virus constructed using reverse genetic techniques could be further developed as a novel genetic engineering vaccine against infectious bronchitis.
Animals ; Cercopithecus aethiops ; Chick Embryo ; Chickens ; Coronavirus Infections ; veterinary ; virology ; Infectious bronchitis virus ; chemistry ; genetics ; growth & development ; metabolism ; Poultry Diseases ; virology ; Protein Structure, Tertiary ; Spike Glycoprotein, Coronavirus ; chemistry ; genetics ; metabolism ; Transfection ; Vero Cells

A HPLC method has been developed in the current investigation for simultaneous determination of three chemical markers of by akangelicin, imperatorin and isoimperatorin in Radix Angelicae Dahuricae. Separation was performed at 30 degrees C. on achromatographic column of Platisil ODS C18 (4.6 mm x 250 mm, 5 μm). The mobile phase was acetonitrile-water. The flow rate was 1.0 mL x min(-1) and the detection wavelength was 254 nm. The results showed that the three chemical markers could be well resolved and that in the selected linear range, all calibration curves of the three chemical markers showed good linearity (R2 ≥ 0.999 8). The recoveries of byakangelicin, imperatorin and isoimperatorin were 100.83%, 100.10% and 103.52%, respectively, and RSD were 1.7%, 0.77% and 0.41% (n = 6), respectively. The data suggested that the developed HPLC method had good reproducibility, robustness, and accuracy, which was suitable for the quality control of Radix Angelicae Dahuricae. Applications of this method showed that the three chemical markers had higher contents in the Bozhou Anhui and Changge Henan than others.
Angelica ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; analysis ; Furocoumarins ; analysis ; Plant Roots ; chemistry ; Quality Control

OBJECTIVE: To investigate the effect of diazepam and modafinil on acute hepatic failure in mice. METHODS: Acute liver failure was induced in male Kunming strain mice by enterocoelia injecting the mice with D-GalN and LPS . The mice in the treatment groups were given corresponding drug 2 h before the administration of D-GalN and LPS, and the mice in the control group were given the same dose of distilled water. The 24-hour survival rate, serum alanine aminotransferase (ALT), and aspartate aminotransferase (AST) levels were compared. Serum levels of TNF-alpha and IL-1 and the levels of SOD, MDA, GR, GSH, NO and NOS in the liver were determined. RESULTS: Treatment with diazepam increased the survival rate and improved liver histological feature. Diazepam inhibited the serum levels of ALT, AST, TNF-alpha and IL-1, and reduced levels of MDA, NO and NOS and increased levels of GR and SOD in the liver. Modafinil decreased liver histological feature, increased the serum levels of ALT, AST, TNF-alpha and IL-1, increased level of MDA, and inhabited levels of SOD and GR in the liver. CONCLUSION Treatment with diazepam may suppress the D-GalN/LPS-induced acute hepatic failure and modafinil may facilitate the acute hepatic failure.
Animals ; Benzhydryl Compounds ; adverse effects ; therapeutic use ; Diazepam ; adverse effects ; therapeutic use ; Galactosamine ; Lipopolysaccharides ; Liver ; pathology ; Liver Failure, Acute ; chemically induced ; drug therapy ; pathology ; Male ; Mice ; Modafinil ; Random Allocation

OBJECTIVETo investigate the application of reverse island auricular composite tissue flap based on the superficial temporal artery for alar defects.

METHODS16 cases with alar defects were treated with reverse island auricular composite tissue flap based on the superficial temporal artery.

RESULTSAll the composite auricular flaps survived with primary healing. The patients were followed up for half a year to four years. The survived composite flaps had a similar color, texture and appearance as normal alar. The cosmetic result was satisfactory.

CONCLUSIONSReverse island auricular composite tissue flap based on the superficial temporal artery is very suitable for alar defects.

Adult ; Ear, External ; transplantation ; Female ; Humans ; Male ; Middle Aged ; Nose Deformities, Acquired ; surgery ; Reconstructive Surgical Procedures ; methods ; Skin Transplantation ; Surgical Flaps ; blood supply ; Temporal Arteries ; surgery ; Young Adult

OBJECTIVETo investigate the injury effect of high frequency electrocoagulation (HFE) by ultrafine-needle on the cutaneous capillary hemangiomas by histopathologic study of cock comb.

METHODS32 male cocks were randomly divided into 4 groups with 8 animals in each group. 3 experimental groups received HFE with different power (6W, 8W, 10W) on the cock comb. Specimens were taken from cock combs for histologic study immediately after treatment and 7, 14, 21, 28 days later.

RESULTSAfter treatment, the cock combs turned pale and followed with scar formation. The shape of combs changed a lot after high power treatment. Immediately after treatment, the capillary number in the experimental groups decreased markedly under light microscope, compared with that in control group (P < 0.05). After that, the capillary number in experimental group increased. 7 days after treatment, the capillary number in experimental groups was still significantly lower than that in control groups (P < 0.01), but not markedly different between the three experiment groups (P > 0.05). Collagen formation also increased after treatment (P < 0.05), which was higher in experimental groups with high power (P < 0.05, correlation coefficient r > 0.5).

CONCLUSIONSHFE with ultrafine needle can effectively decrease the capillary number in cock comb. The 6W treatment is mini-invasive and will not destroy the comb shape with good cosmetic results. The technique may be used for the treatment of cutaneous capillary hemangioma.

Animals ; Capillaries ; pathology ; surgery ; Chickens ; Disease Models, Animal ; Electrocoagulation ; methods ; Hemangioma, Capillary ; pathology ; surgery ; Male ; Random Allocation ; Skin Neoplasms ; pathology ; surgery

OBJECTIVETo investigate the therapeutic effect of scalp flaps pedicled with superficial temporal vessel and hair removal for reconstruction of facial skin defects.

METHODSFrom Jan. 2010 to Dec. 2011, 6 cases with large facial skin defects were treated with scalp flaps pedicled with superficial temporal vessel and hair removal. At the first stage, the skin expanders were implanted subcutaneously at the homolateral side according to the defect size. After the expansion was finished, the expanded flaps pedicled with superficial temporal vessel were used to reconstruct the facial skin defects at the second stage. 2 weeks after operation, hair removal was performed to remove the hair on flaps. 4-6 treatments were needed.

RESULTSSeven flaps in 6 cases were survived completely. The wounds at donor sites were closed directly. The effect of hair removal was reliable. The patients were followed up for 6 months to 1 year. The flap color, texture and thickness had a good match with surrounding skin tissue.

CONCLUSIONSIt is a good option to reconstruct facial skin defects with scalp flaps pedicled with superficial temporal vessel and hair removal.

Adolescent ; Face ; surgery ; Hair Removal ; Humans ; Reconstructive Surgical Procedures ; Scalp ; transplantation ; Surgical Flaps ; blood supply ; transplantation

OBJECTIVETo study the chemical constituents in hairy roots of Cassia obtusifolia.

METHODThe hairy roots of C. obtusifolia were induced with Agrobacterium rhizogenes LBA9402 from cotyledons and cultured in MSO liquid medium. The compounds were isolated by silica gel, polyamide and Sephadex LH-20 column chromatography, and the structures were elucidated by employing chemical and spectral methods

RESULTEight compounds were isolated from the ethyl acetate fraction of 95% EtOH extract of the transformed roots of C. obtusifolia. They are betulinic acid, chrysophanol, physcion, stigmasterol, 1-hydroxy-7-methoxy-3-methyl-anthraquinone, 8-O-methylchrysophanol, 1-O-methylchrysophanol and aloe-emodin, and aloe-emodin was isolated from the hairy roots of C. obtusifolia for the first time.

CONCLUSIONThe hairy roots of C. obtusifolia have the ability to synthesize the similar chemical constituents as the original plants.

Anthraquinones ; chemistry ; isolation & purification ; Cassia ; chemistry ; growth & development ; microbiology ; Cotyledon ; growth & development ; microbiology ; Culture Media ; Emodin ; analogs & derivatives ; chemistry ; isolation & purification ; Plant Roots ; chemistry ; growth & development ; microbiology ; Plants, Medicinal ; chemistry ; growth & development ; microbiology ; Rhizobium ; Tissue Culture Techniques ; methods ; Triterpenes ; chemistry ; isolation & purification

OBJECTIVETo study the induction of hairy roots and anthraquinone production in Rheum palmatum.

METHODLeaf blades, hypocotyls and petioles of R. palmatum were infected by Agrobacterium rhizogenes LBA9402 and A. rhizogenes R1601, respectively.

RESULTHairy roots were induced by two strains of A. rhizogenes, and LBA9402 showed stronger infective ability than R1601. Three clones of hairy roots were cultured. DH7a was induced by R1601, DH5a and DH5c were induced by LBA9402. DH7a grew faster than DH5a and DH5c, and all of them grew faster obviously than normal root (NOR). There were significant differences in anthraquinone composition and content among four kinds of roots. Emodin physcion and chrysophanol were predominant anthraquinone in DH5a, DH5c and NOR respectivly. Aloe-emodin content was the lowest in all root cultures compared with other four anthraquinones.

CONCLUSIONThe in vitro culture system of the established hairy roots laid a foundation for mass production of anthraquinone by hairy root culture.

Anthraquinones ; analysis ; Emodin ; analogs & derivatives ; analysis ; Plant Roots ; chemistry ; growth & development ; microbiology ; Plants, Medicinal ; chemistry ; growth & development ; microbiology ; Rheum ; chemistry ; growth & development ; microbiology ; Rhizobium ; physiology ; Tissue Culture Techniques ; methods

OBJECTIVETo study the feasibility of regenerating a whole menisci using poly-(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) scaffolds loaded with meniscal cells in rabbits undergoing total meniscectomy, and to explore its protective effect on cartilage degeneration.

METHODSA solvent casting and particulate leaching technique was employed to fabricate biodegradable PHBV scaffolds into a meniscal shape. The proliferated meniscal cells were seeded onto the polymer scaffolds, transplanted into rabbit knee joints whose lateral menisci had been removed. Eight to 18 weeks after transplantation, the rege- nerated neomenisci were evaluated by gross and histological observations. Cartilage degeneration was assessed by Mankin score.

RESULTSEighteen weeks after transplantation, the implants formed neomenisci. Hematoxylin and eosin (HE) staining of the neomenisci sections revealed regeneration of fibrocartilage. Type I collagen in the neomenisci was also proved similar to normal meniscal tissue by immunohistochemical analysis and Sirius scarlet trinitrophenol staining. Articular cartilage degeneration was observed 8 weeks after implantation. It was less severe as compared with that in total meniscectomy controls and no further degeneration was observed at 18 weeks. At that time, the regenerated neomenisci strongly resembled normal meniscal fibrocartilage in gross and histological appearance, and its mechanical property was also close to that of normal meniscus.

CONCLUSIONSThe present study demonstrates the feasibility of tissue-engineering a whole meniscal structure in total meniscectomy rabbit models using biodegradable PHBV scaffolds together with cultured allogeneic meniscal cells. Cartilage degeneration is decreased. But long-term in vivo investigations on the histological structure and cartilage degeneration of the neomenisci regenerated by this method are still necessary to determine the clinical potential of this tissue engineering avenue.

Animals ; Cartilage, Articular ; Cells, Cultured ; Knee Joint ; Menisci, Tibial ; Polymers ; Regeneration ; Tissue Engineering

Objective To determine the clinical features,neurophysiological characteristics and cervical magnetic resonance imaging of sensory neumnopathy,and to describe the pathology of skin nerve, sural nerve and spinal dorsal columns.Methods Two patients who died from sensory neuron disease (SND)after infection of digestive tract were discussed including clinical features and ancillary tests which included neurophysiology and pathology of peripheral nerve and spinal dorsal columns.Associated documents are reviewed.Results Early ataxia,widespread sensory symptoms and global loss of deep tendon reflex were the distinctive signs of SND,which was characterized by non-length-dependent abnormalities of sensory nerve action potentials,a hallmark of ganglionopathies.The second patient showed normal cervical magnetic resonance imaging possibly because of short course of disease,while diffuse hyperintensity in the spinal posterior columns of SND was reported.Demyelination of spinal posterior columns and loss of mostly large diameter nerve fibers without regeneration clusters were the main pathological features.Conclusions The distinctive clinical features and neurophysiological characteristics of SND indicate that peripheral sensory nerve fibers are widely damaged.Pathology of spinal posterior columns confirm that central sensory pathway are impaired which allow the localization of the pathologic site to the dorsal root ganglion neurons.Cervical spinal MRI of SND are possibly normal at early phase.