BACKGROUND: There are no reports about adult degenerative disc cell culture model currently. This establishment of disc cell culture is the cellular basis of intervertebral disc tissue engineering. OBJECTIVE: To establish cell culture models from human degenerative intervertebral disc, and to settle the cytology base of disc tissue engineering. DESIGN AND SETTING: The controlled observational experiment with regard to cytology was performed in Shandong Institute of Orthopaedics and Traumatology. MATERIALS: Five degenerative intervertebral disc patients were recruited from Department of Orthopaedics, Affiliated Hospital of Qingdao University Medical College, including one case in L2-3, three cases in L4-5 and one case in L5-S1. METHODS: Five samples were taken from the operational disectomy for lumbar disc herniation, and these samples were cultured using enzymatic digestion cell culture and tissue cell culture method differently. All the samples were cultured with HAMF12 medium with the addition of 10% and 20% fetal bovine serum. The morphology of the cultured cell was observed with Giernsa staining and transmission electron microscopy respectively. MAIN OUTCOME MEASURES: ①Cellular growth; ②Morphology of the cultured cells;③Ultrastructure Of the cultured cells. RESULTS: By means of enzymatic digestion cell culture and tissue cell culture method, numerous degenerative lumbar disc cells were obtained and successfully subcultured. The cell secretions around the cultured cells may influence the growth of these cells. Destroying the secretions, the cultured cells could be highly proliferated. The degenerative intervertebral disc cells were proliferated vigorously in HAMF12 medium with 20% fetal bovine serum compared with that with 10% fetal bovine serum. The notochordal cell was observed in all the specimens. CONCLUSION: The adult degenerative disc can be used to obtain the required cell for tissue engineering.

Objective To compare the effects of fresh amniotic membrane (FAM), glycerol-preserved amniotic membrane (GPAM), alcohol-preserved amniotic membrane (APAM) and amniotic membrane preserved under vacuum drying condition (VXAM) on rabbit corneal alkali burn and to observe the outcomes of the grafted amniotic membrane. Methods Thirty-six New Zealand rabbits were used to establish corneal alkali burn models and then divided into 4 groups according different preserved amniotic membrane they received (FAM, GPAM, VXAM and APAM), with 9 rabbits in each group. After transplantation, we observed the changes of amniotic membrane graft, corneal opacity, corneal neovascularization (CNV) with slit lamp microscopy. The amniotic membrane graft was examined by HE staining 10 d, 14 d and 1 month after transplantation. Results One month later, FAM graft, GPAM graft and VXAM graft showed better curative effect than APAM graft. Ten days after transplantation, the amniotic membrane graft began to dissolve and desquamate, and disappeared completely till 14 d after transplantation. Amniotic membrane tissues were not found under light microscope in any rabbit. FAM graft dissolved most quickly, and APAM graft was the most slowly. The dissolution of GPAM graft and VXAM graft had no significant difference. Conclusion After amniotic membrane transplantation, it dissolves and sheds instead of fusing with recipient tissues. Corneal epithelium grows under amniotic membrane graft. Amniotic membrane graft transplantation should be called as amniotic membrane patching.

AIM:Recently,amnion transplantation has been one of effective methods for curing corneal alkali burn,but its biomechanism is unclear.This study was to discuss the efficiency of different kinds of amnion on rabbit corneal alkali burn and to find the bioactive functions of the amnion.METHODS:The experiment was carried out in the Central Laboratory of Experimental Animals at Chongqing University of Medical Sciences between January and July in 2004.All the disposals were in accordance with the ethical standard of animals.①Placenta from healthy puerperants underwent uterine-incision delivery were offered by Department of Obstetrics in the First Affiliated Hospital of Chongqing University of Medical Sciences.A total of 45 New Zealand rabbits were used for corneal alkali burn models and divided into fresh amniotic membrane(FAM),glycerin preserved amniotic membrane(GPAM),vacuum xeransis amniotic membrane(VXAM),alcolhol preserved amniotic membrane(APAM) and control groups,every group had 9 rabbits.②The FAM was prepared within 24 hours,and the preserved amnion were transplanted after 1 month.③The corneal opacity and corneal neovascularization were observed everyday during days 2-21 after operation and once every three days after days 21;corneal fluorescein staining was conducted to observe the repair of corneal epithelium on days 14,30,60 after operation;2 weeks,1 month and 2 months after operation,we did histopathological examination and scanning electron microscope observation.RESULTS:Forty-three rabbits were involved in the result analysis.①There were no significant differences in the corneal opacity and repair velocity of corneal epithelium among FAM,GPAM and VXAM,which were remarkably superior to APAM and controls.APAM group and control group had not distinct difference.②FAM had better effects on inhibiting corneal neovascularization than GPAM and VXAM.CONCLUSION:After transplantation,VXAM,GPAM and FAM can all accelerate corneal epithelial healing and lighten inflammation.FAM containing more bioactive materials,plays best.

Background Our previous study showed that curcumin suppresses the proliferation of human lens epithelial cells (LECs) in vitro and has an influence on the signal transduction of cyclic adenosine monophosphate (cAMP) and cyclic guanosinc monophosphate (cGMP).Actually,the regulation for biological behavior of LECs in vivo is complex.Objective This study was to investigate the changes of signal transduction of protein kinase (PK) in inhibition of curcumin on human LECs proliferation.Methods HLE-B3 was cultured and then divided into the blank control group,recombinant human basic fibroblast growth factor (rhbFGF) group and rhbFGF+ curcumin group.rhbFGF of 10 ng/ml was added in the medium in the rhbFGF group,and 20 mg/L curcumin was added into rhbFGF-induced cell medium for 24 hours in the rhbFGF+ curcumin group.The expression rates of PKA,PKC,PKG and calmodulin (CaM) in the cells were assayed using flow cytometry.Results The expression rates of PKA protein were (46.847± 1.673) ％,(33.250 ± 2.242) ％ and (71.645 ±2.432) ％ in the blank control group,rhbFGF group and the rhbFGF+ curcumin group,respectively,and the expression rate of PKA protein was significantly reduced in the rhbFGF group compared with the blank control group (t =11.904,P＜0.01),but the expression rate of PKA protein in the rhbFGF+ curcumin group was significnatly higher than that in the rhbFGF group (t=28.430,P＜0.01).In the blank control group,rhbFGF group and the rhbFGF+ curcumin group,the expression rates of PKC protein in the cells were (35.575± 1.937) ％,(50.652±2.068) ％ and (27.662t4.481) ％,those of PKG protein were (63.838±0.486) ％,(86.562 ± 1.325) ％ and (40.733 ± 2.175) ％,while those of CaM protein were (67.408± 1.391)％,(83.887±3.499)％ and (53.785 ± 1.942)％,the expression rates of PKC,PKG and CaM were significantly lower in the rhbFGF group in comparison with the blank control group (all at P＜0.01),and those in the rhbFGF+ curcumin group were significantly declined in comparison with the rhbFGF group (all at P＜0.01).Conclusions Suppression of curcumin on HLE-B3 proliferation probably is associated with the up-regulation of PKA expression and down-regulation of PKC,PKG and CaM expression in the cells.

Objective To suppress the proliferation of lens epithelial cells (LECs) is a primary goal in prevention of after cataract. Recent study demonstrated an effective inhibition of elemene(Ele) on tumor cells. Present study was to investigate the effects of Ele on proliferation and cell cycle of human LECs B3 (HLE-B3). Methods Recombinant human basic fibroblast growth factor(rhbFGF) was utilized to induce proliferation of HLE-B3. Proliferative HLE-B3 was incubated with 80 mg/L Ele in CO_2 incubator for 24 hours. Then the inhibitory effects of Ele on proliferation of HLE-B3 was evaluated by methyl thiazolyl tetrazolium(MTT). The effect of Ele on HLE-B3 morphology was observed under the optical microscope. The effect of Ele on HLE-B3 cell cycle was analyzed by flow cytometer(FCM). Results MTT test showed that the optical density (OD) value of rhbFGF group was remarkably higher than that of control group(0. 599 0 ± 0. 053 1 versus 0. 409 1 ± 0. 042 2) (P ＜ 0. 01), and that of Ele group(0. 450 0 ±0. 061 4) was obviously declined in comparison to rhbFGF group(P ＜0. 01) . The inhibitory rate of Ele was 24.90 %. In proliferation group, the number of HLE-B3 was increased with the normal cell structure and abundant cytoplasm under the optical microscope. However, in Ele group, the number of HLE-B3 was evidently decreased with less cytoplasm, undistinguished cell structure, condensed and aggregated nucleuses. The result of flow cytometer showed that the percentage of HLE-B3 in G_1 phase in rhbFGF group was 42. 062% ± 1. 270% and that in control group was 46. 422% ±3. 765% with a significant difference(P ＜ 0. 05). HLE-B3 in G_1 phase in Ele group (60. 665% ±2.069%) was evidently increased in comparison with rhbFGF group (P ＜ 0. 01). HLE-B3 in S phase in rhbFGF group compared with control group was increased (51.647% ±1.123% versus 31.842% ± 2. 798%) (P ＜ 0. 01), but that in S phase in Ele group(30. 222% ±3.429%) was lower than rhbFGF group (P ＜ 0. 01). HLE-B3 in G_2 phase in rhbFGF group was decreased in comparison with control group (6. 288% ± 0. 966% versus 21. 735% ± 3. 806%, P ＜ 0. 01), and that in G_2 phase in Ele group (9. 112% ± 1. 659%) compared with proliferation group was increased (P ＜ 0. 01). Conclusion Ele could alter the cell cycle of HLE-B3 and effectively inhibit the HLE-B3 proliferation induced by rhbFGF. Ele may be a reliable and effective drug for prevention and treatment of after cataract.

Objective To assess polymerase chain reaction(PCR)combined with restriction fragment length polymorphism(RFLP)and gene sequencing technologies in the detection and typing of HPV DNA.Methods Tissue specimens were collected from skin diseases and venereal disease in perianal or genitals.PCR was performed with HPV DNA general primers(MY09/11)in tissue samples. Positive fragments of HPV DNA were purified and digested by restriction enzymes.The digested fragments were typed by po]yacrylamide gel electrophoresis(PAGE).The Resultswere verified by direct sequencing.Results In 50 clinical samples there were 35 HPV DNA positive,including 26 from patients with condyloma acuminatum,8 from patients with bowenoid papulosis,and 1 from patients with squamous cell carcinoma.In HPV DNA positive samples,19 were HPV6,3 were HPV11,8 were HPV16,4 were HPV6 and HPV 11,and I was HPV62.Sequencing Resultswere in accordance with the PCR-RFLP Results .Conclusion PCRRFLP method is effective in the detection and typing of HPV DNA.

Wearable spectral sensors are integrated smart devices consisting of flexible substrates,sensing and detection units,and analysis units.These sensors can track chemicals in human fluids including sweat,transdermal gases,wound secretions,subcutaneous mesenchymal fluids,tears,exhaled gases,and saliva quickly and non-invasively.Their application is crucial in drug analysis,disease diagnosis,and health monitoring.This paper summarized the literature studies of wearable colorimetric,fluorescence and surface-enhanced Raman scattering sensors since 2016,and reviewed the advancements in wearable spectroscopic sensors,focusing on their composition and applications in epidermal,ocular,and oral forms.Additionally,the future challenges and opportunities of wearable spectroscopic sensors were discussed,providing a reference for the design and development of wearable spectroscopic sensors.

This study was intended to look for anti-HIV chemical constituents of aerial parts of Caragana rosea Turcz. Column chromatographic technique was used for the isolation and purification of constituents of Caragana rosea under the guide of anti-HIV assay. The structures were established on the basis of physical and chemical properties and spectroscopic data. Five compounds were obtained from the EtOAc fraction of aerial parts of Caragana rosea and identified as myricetin (1), mearnsetin (2), p-hydroxy cinnamic acid (3), cararosinol A (4) and cararosinol B (5). At the same time, one possible transformation route between cararosinol B and kobophenol A, another resveratrol tetramer isolated from this plant previously, was proposed. Compounds 4, 5 are new resveratrol tetramers, compounds 1 -3 were isolated from this plant for the first time. All compounds showed no activities in an in vitro assay against HIV-1.
Anti-HIV Agents ; chemistry ; isolation & purification ; pharmacology ; Benzofurans ; chemistry ; isolation & purification ; pharmacology ; Caragana ; chemistry ; Coumaric Acids ; chemistry ; isolation & purification ; pharmacology ; Flavonoids ; chemistry ; isolation & purification ; pharmacology ; HIV-1 ; drug effects ; Molecular Structure ; Plant Components, Aerial ; chemistry ; Plants, Medicinal ; chemistry ; Propionates ; Stilbenes ; chemistry ; isolation & purification ; pharmacology

Cases of high-temperature requirement A serine peptidase 1(HTRA1)gene heterozygous mutation associated cerebral small vessel disease are relatively rare.Early and timely diagnosis and treatment can improve prognosis.The authors reported a 37 years old male patient admitted in Department of Neurology,the Second Affiliated Hospital of Nanchang University,whose initial symptom was transient right limb weakness,imaging suggested white matter lesions and the gene screening showed HTRA1(c.854 C＞T/p.Pro285Leu)heterozygous mutation.A family survey has been conducted and the characteristics of patients in this family are as follows:they present with ischemic cerebrovascular disease,coexist with cervical or lumbar disc herniation,male patients have hair loss,some patients have cognitive dysfunction,men tend to develop the disease at an earlier age than women,and the onset age is progressively earlier from generation to generation.Therefore,for young ischemic cerebrovascular disease patients with hair loss,cognitive dysfunction,cervical or lumbar disc herniation,and obvious white matter lesions on imaging,especially those without common risk factors for cerebrovascular disease,a family history should be inquired and genetic testing should be performed to screen for HTRA1 mutations.

OBJECTIVETo investigate the protective effects of the natural medicinal monomer isopsoralen (ISR) with estrogenic activity against oxidative damage in human lens epithelial cells B3 (HLE-B3) caused by hydrogen peroxide (H(2)O(2)) and to pursue the possible mitochondrial proteomic regularity of the protective effects.

METHODSHLE-B3 cells were treated with H(2)O(2) (300 μ mol/L), β-estradiol (E(2): 10(-8) mol/L) and H(2)O(2), ISR (10(-5) mol/L) and H(2)O(2), or left untreated. Altered expressions of all mitochondrial proteins were analyzed by protein array and surfaceenhanced laser desorption ionization time of flight mass spectrometry (SELDI-TOF-MS). The mass/charge (m/z) ratios of each peak were tested by the Kruskal-Wallis rank sum test, and the protein peak value of the m/z ratio for each treatment by pair comparison was analyzed with the Nemenyi test.

RESULTSH(2)O(2) up-regulated the expressions of two protein spots (with m/z of 6532 and 6809). E(2) mitigated the oxidative damage, and the expression of one protein spot (m/z 6532) was down-regulated. In contrast, ISR down-regulated both of protein spots (m/z 6532 and 6809).

CONCLUSIONSISR could effectively inhibit H(2)O(2)-induced oxidative damage in HLE-B3 cells. The protein spot at m/z of 6532 might be the target spot of ISR against oxidative damage induced by H(2)O(2).

Cell Line ; Epithelial Cells ; drug effects ; metabolism ; pathology ; Estradiol ; pharmacology ; Furocoumarins ; pharmacology ; Humans ; Hydrogen Peroxide ; toxicity ; Lens, Crystalline ; pathology ; Mitochondria ; metabolism ; Oxidation-Reduction ; drug effects ; Oxidative Stress ; drug effects ; Protective Agents ; pharmacology ; Proteome ; metabolism ; Proteomics ; methods