Effect of insulin on the secretion of VEGF and its receptor in serum-free cultured bovine thoracic aortic endothelial cells

AIM: This study was designed to investigate the secretion of VEGF and its receptor (flt-1 or flk-1/KDR) protein by cultured bovine thoracic aortic endothelial cells treated with various insulin concentrations. METHODS: Endothelial cells was isolated from bovine thoracic aorta, and cultured in serum-free medium, then incubated with different insulin concentrations (30 mU/L, 300 mU/L, 3 000 mU/L). The level of VEGF and its receptor (flt-1 or flk-1/KDR) protein were detected by immunohistochemical staining. RESULTS: As compared with no insulin group, the expression of VEGF protein in low insulin concentration (30 mU/L and 300 mU/L) groups were significantly increased (P＜0.01). The expression of VEGF protein in high insulin concentration (3 000 mU/L) group was significantly decreased (P＜0.05). Howerer, no difference of the expression of VEGF receptor (flt-1 or flk-1/KDR) protein among all groups (P＞0.05) was observed. CONCLUSION: Low concentration insulin up-regulates the VEGF protein expression while high concentration insulin down-regulates the VEGF protein expression in bovine thoracic aortic endothelial cells, but insulin had no directly effect on the VEGF receptor (flt-1 or flk-1/KDR) protein expression in bovine thoracic aortic endothelial cells.     

Effect of EGFP gene transfection on the cell cycle distribution of primary cultured human chondrocytes

AIM: To investigate the effect of enhanced green fluorescence protein (EGFP) gene transfection on the cell cycle distribution of primary cultured human chondrocytes in order to establish a tracking method of cultured human nasoseptal chondrocytes. METHODS: pEGFP-N1 plasmid was amplified in E.coli, and purified by high purity kit. Primary cultured human chondrocytes,which were initially obtained from the nasoseptal cartilage, were cultured in vitro and transferred with pEGFP-N1 by means of electroporation with Amaxa nucleofector device. Transfering process and transient expression were evaluated by laser scanning confocal microscope (LSCM), the transfer efficiency and the cell cycle distribution were evaluated by flow cytometry. RESULTS: There was significant expression of EGFP at 24 h after transferring. The transfection efficiency of pEGFP-N1 into primary cultured human chondrocytes reached 35.37％ at 48 h. It didn't affect the process of cell adherance and had no effect on the cell cycle distribution. CONCLUSION: Primary cultured human chondrocytes, which were transfected with pEGFP, are alive in vitro, and the transferring process doesn't affect the cell cycle distribution. These results suggest that pEGFP-N1 is an ideal transient expression vector for primary cultured human chondrocytes and it might be a well tracer in construction tissue engineered cartilage.     

Osmolarity,cell volume and proliferation in nasopharyngeal carcinoma cells

AIM: To investigate the relationship between osmolarity, cell volume and cell proliferation in nasopharyngeal carcinoma cells. METHODS: MTT method was applied to detect the proliferation ability of the poorly-differentiated nasopharyngeal carcinoma cell (CNE-2Z) under various osmolarity conditions. The flow cytometry was used to analyse cell cycle distribution. Cell volume was obtained by the image analysis of living cells and cell viability was determined by the trypan blue assay. RESULTS: Cultivation of cells under the hypertonic conditions of 370 and 440 mOsmol/L increased cell volume by 8.7% and 27.8% and facilitated cell proliferation by 22.2% and 33.9%, respectively. However, hypotonic incubation of cells with osmolarity of 160 and 230 mOsmol/L decreased cell volume by 12.8% and 4.1% and inhibited cell proliferation by 34.0% and 15.6%, respectively. Cell volume was positively correlated with cell proliferation rate. Long-term cultivation of cells under anisotonic conditions did not significantly alter cell cycle distribution, but hypotonic cultivation decreased cell viability. CONCLUSION: Proliferation of nasopharyngeal carcinoma cells was closely correlated with the osmolarity of culture medium and cell volume. Hypotonic cultivation may inhibit cell proliferation by decreasing cell volume to facilitate cell death mechanisms.     

Effects of component of some Chinese herbs on proliferation of human umbilical vein endothelial cells in vitro

AIM: To observe the effects of some component of Chinese herbs for external use on proliferation of human umbilical vein endothelial cells (HUVEC) and investigate the mechanism of promoting tissue repair. METHODS: The method of MTT was used to examine the effects of Rg1, Rh1, perlolyrine, cinnamyl aldehyde, muscone, astragaluspolysaccharin (APS), velver antler polypeptide (VAP) and soluble extract of boswellia carterii birdw (BCB) on proliferation of HUVEC. RESULTS: APS did not promote proliferation of HUVEC at 9.75 mg/L-2.5 g/L; Rh1 promoted proliferation of HUVEC at 1.94 mg/L-0.5 g/L (P<0.05 or P＜0.01), and Rg1 inhibited proliferation of HUVEC at 31 mg/L (P<0.05); VAP promoted proliferation of HUVEC at 1 mg/L-0.5 g/L with optimal dose of 10 mg/L (P<0.01), Cinnamyl aldehyde promoted proliferation of HUVEC at 2 g/L(P＜0. 05); Muscone and soluble extract of BCB inhibited proliferation of HUVEC at 1 g/L, 0.5-2.5 kg/L(P＜0. 01), respectively; Perlolyrine inhibited proliferation of HUVEC at 0.125 g/L-0.5 g/L(P＜0. 01). CONCLUSION: The external herbs for supplementing Qi and warming Yang can promote HUVEC proliferation and improve angiogenesis during tissue repair. The external herbs for promoting blood circulation and accelerating capillary movement may have influence upon other stages of tissue repair.     

Recent advances in molecular mechanisms of oncosis

Oncosis is another form of cell death, which is different from apoptosis. The review will discuss the recent advances of oncosis on pathological morphology, nuclear biochemical changes and molecular mechanisms.     

山羊胎期肺发育的形态学研究

对3～22周龄山羊胎儿肺进行了肉眼、光镜和透射电镜观察，结果表明：1．7～22周龄山羊胎儿肺的外部形态与胎龄无关，肺的外部形态以左二右四叶者为多见．肺的叶间裂和右肺副裂常不完整，以浆膜、肺组织或混合性组织(肺组织及浆膜)融合；2．山羊胎儿肺的发育分为5个时期：胚胎期(3～5周)肺芽分支形成主支气管，主支气管长度不断增长并萌芽出叶支气管，均衬以假复层柱状上皮。腺状期(6～12周)以支气管树发育为主，小支气管衬以假复层和／或单层柱状上皮；终蕾呈腺状，上皮细胞由假复层柱状逐渐变为单层柱状，胞核向细胞顶端移行；终蕾上皮细胞游离面可见短小的微绒毛；线粒体、粗面内质网及核糖体随着胎龄增加而逐渐增多，它们均位于细胞顶部。小管期(13～14周)以呼吸部发育为主，原始肺泡开始形成，呼吸性细支气管衬以未分化的立方上皮；终蕾腺状结构逐渐消失，终蕾上皮细胞由高矮不等的单层柱状上皮逐渐演变为立方形的原始肺泡上皮；细胞游离面可见较多的微绒毛，胞质内线粒体、粗面内质网及核糖体较发达。囊状期(第15周)呼吸部发育显著，肺内细支气管及其末端呈现出“充气”状态；部分原始肺泡上皮细胞分化为扁平的肺泡Ⅰ型细胞和立方形的肺泡Ⅱ型细胞；Ⅱ型细胞内出现嗜锇小体。肺泡期(16～22周)以肺泡的形成和分化为主，更多的肺泡上皮分化为扁平的肺泡Ⅰ型细胞和立方形的肺泡Ⅱ型细胞。此期，毛细血管内皮与部分肺泡上皮贴近，可将肺泡上皮细胞区分为3种：Ⅰ型细胞，呈矮柱状或椭圆形，胞质中有较明显的核糖体、扩张内质网及变性线粒体；形成了由Ⅰ型细胞一基膜一内皮细胞组成的气血屏障。Ⅱ型细胞，胞质内含丰富的嗜锇板层小体和核糖体，内质网扩张呈大小不一的泡状，多泡体出现，线粒体膨大变性，细胞游离面可见少数微绒毛。Ⅲ型细胞，为未分化细胞，呈立方形，胞体较小，胞核相对较大，呈圆或椭圆形，胞质少，呈带状．电子密度低，细胞器少。     

Morphofunctional description of mucous cells in the gills of the Arapaimidae Arapaima gigas (Cuvier) during its development

The gill structure of the Amazonian fish Arapaima gigas (Cuvier 1829) shows ontogenetic changes during development, particularly due the transition from the aquatic to the obligatory air breathing mode of respiration. However, three main cell types can be found in the gills: mitochondrial rich cells, pavement cells and mucous cells (MCs). The MCs are involved in the secretory pathway. The functions of the secreted molecules include mechanical protection of epithelia, protection against parasites and bacterial infection, and role on ion regulation. In this study, we analysed mucous cell location and mucous cell type, based on pH, during the development of A. gigas. Using samples obtained from the environment, gills were collected and fixed in buffered solution. Histological techniques for the identification of MCs were performed Alcian Blue (AB) and periodic acid‐Schiff (PAS). The results showed the presence of PAS+ and AB+ cells in the whole filament in all examined fish. In animals less than 50 g, few MCs were present, and no differences were observed in AB+ and PAS+ cells. In animals weighing close to 500 g, more PAS+ cells than AB+ cells were observed, and in animals that weighed more than 1,000 g, more AB+ cells than PAS+ cells were observed. These observations may be a result of the ontogenetic changes in the gill epithelia, which can change the osmorespiratory compromise in ion regulation functions as well the glycosaminoglycans secreted by PAS cells, which in large animals can play a role in the protection against parasites and bacterial infection.     

The evolution and application of enzymes in the animal feed industry: the role of data interpretation

ABSTRACT

1. Enzymes have been used commercially for nearly 40 years and save significant costs through sparing of expensive nutrients but the mechanism by which this is achieved is still debated.

2. The research focused on non-starch polysaccharidase (NSPase) enzymes is used as an example of where greater progress could have been made if the details of the work had been described more fully and the analysis of the data generated had been broader in scope and more critical.

3. Lack of standardisation of the details presented in the materials and methods has been identified as a significant barrier to meaningful retrospective analysis and thus limits advances in the understanding of the mode of action of these enzymes.

4. The identity of the enzyme employed and its activity is often lacking, and more importantly the purity is rarely disclosed. Contaminant activities which are neither listed nor assayed could play a significant role in the responses observed.

5. The dose optimum of most enzymes is often considerably higher than that employed in most studies. Thus studies claiming synergy between two ‘activities’ should ensure that the response is not related to each enzyme simply augmenting the dose of just one activity in the finished feed. This is a common problem, and coupled with the lack of factorial experiments to justify the presence of each enzyme in a multi-enzyme product, it is not surprising that there is still debate as to whether single or multi-enzymes are best suited poultry rations.

6. The three proposed mechanisms for NSPases (viscosity, cell wall and prebiotic) are discussed, and along with their strengths and weaknesses it is suggested that a re-evaluation of each is needed. Viscosity may have to be re-evaluated as being a function not only of the cereal being fed, but of the age of the animal as well. The cell wall theory as described is poorly modelled in vitro and hence the validity of these data is questioned. The prebiotic theory may need significant modification as it appears that the quantities of oligomers produced are insufficient to generate the additional volatile fatty acids (VFA)’s reported. It is likely that all three mechanisms play a role in the responses observed, but the prebiotic mechanism probably plays by far the most important part in low viscosity diets.

7. Future research would be improved if it considered all potential mechanisms when designing a trial. Significant failings are apparent as a result of adherence to tenets in explanation of the results. Most importantly, it should be emphasised that a hypothesis is there to be tested, not defended.     

Effects of in ovo feeding of L‐arginine on the development of digestive organs,intestinal function and post‐hatch performance of broiler embryos and hatchlings

This study was to investigate the effects of in ovo feeding (IOF) L‐arginine (Arg) solution on the development of digestive organs, the duodenal mucosa of broiler embryos and hatchlings, and the growth performance of chicks during the first week post‐hatch. A total of 720 fertilized eggs with similar weight were randomly allocated to three groups, consisting of eight replicates of 30 eggs each. Three treatments were arranged as non‐injected control, diluent‐injected (0.75% NaCl solution) group and Arg solution‐injected group containing 1% Arg, dissolved in diluent. At 17.5 days of incubation, 0.6 ml of IOF solution was injected into amniotic fluid of each egg of injected groups. Results showed IOF of Arg solution increased (p < .05) the chick embryo weight at 19 days of incubation; the body weight gain of post‐hatch broilers during 1–7 days; the weights of liver, pancreas, proventriculus and gizzard; the concentrations of duodenal ghrelin, vasoactive intestinal peptide and glucagon‐like peptide 2; and the duodenum mucosal enzyme activities of alkaline phosphatase, maltase, sucrase and inducible nitric oxide synthase of 7‐day‐old post‐hatch broilers compared with other groups. The IOF of Arg solution also increased (p < .05) the villus height (VH) and the ratio of VH to crypt depth (CD) and decreased (p < .05) the CD in duodenum of broiler embryos and post‐hatch hatchlings, except for the CD at 19 days of incubation. In conclusion, IOF of 1% Arg solution into the amnion at 17.5 days of incubation could improve the development of digestive organs, the duodenal morphology, the releasing of gastrointestinal hormones and mucosal enzyme activities of broiler embryos and hatchlings and finally the growth performance of chicks during the first week post‐hatch. Therefore, IOF of appropriate Arg solution could be an effective technology for regulating early nutrition supply and subsequent growth development in poultry industry.