胡萝卜直根诱导愈伤组织实验研究

目的：探讨胡萝卜直根诱导愈伤组织的最佳条件.方法：以新鲜胡萝卜直根为外植体,将直根皮层、形成层及中轴3个部位分别置于MS（培养基）＋2,4-D（2,4-二氯苯氧乙酸）＋6-BA（6-苄基腺嘌呤）与MS＋NAA（α-萘乙酸）＋6-BA两类培养基中,观察3种外植体愈伤组织的生长情况及诱导率.结果：直根皮层外植体在这两类培养基中均未长出愈伤组织,即愈伤组织诱导率为0;直根形成层及中轴外植体在这两类培养基中均长出愈伤组织,即愈伤组织诱导率为100％,其中在MS＋ NAA＋ 6-BA培养基中愈伤组织质地疏松、颜色浅黄鲜亮、生长旺盛、增殖快速,在MS＋2,4-D＋ 6-BA培养基中愈伤组织质地紧密、颜色深黄、增殖缓慢.结论：诱导疏松型胡萝卜愈伤组织的最佳外植体是胡萝卜直根形成层及中轴,最佳培养基类型是MS＋ NAA＋ 6-BA.     

Experimental superficial candidiasis on tissue models

Candida species are common pathogens causing superficial mycoses primarily affecting the mucosa and the skin in humans. Crucial steps during pathogenesis of superficial candidiasis comprise fungal adhesion, colonisation and subsequent penetration of the respective tissues. Exploring these pathological events and perhaps fungal and tissue responses towards drug treatment is imperative in the management of this infection. Unfortunately, pathological biopsies of superficial candidiasis do not exhibit the early changes in the host–pathogen interaction as the tissues are already invaded by the fungi. In vivo experimental assessments of pathological processes of superficial candidiasis are also limited because of the difficulties in providing reproducible and comparable conditions in the host environment. Conversely, in vitro models have helped studying fungal–host interactions under more defined and controlled conditions. Some common in vitro models used to simulate superficial candidiasis are chick chorioallantoic membrane, mucosal explants and single layer or multiple layer cell cultures. Interestingly, these experimental approaches share advantages as well as disadvantages when compared with in vivo conditions. Hence, this review intends to discuss about the experimental superficial candidiasis produced in various tissue models and their advantages as well as disadvantages with a particular reference to further improvement of validity and reliability of such experiments.     

Depression‐like behaviour in mice is associated with disrupted circadian rhythms in nucleus accumbens and periaqueductal grey

An association between circadian rhythms and mood regulation is well established, and disturbed circadian clocks are believed to contribute to the development of mood disorders, including major depressive disorder. The circadian system is coordinated by the suprachiasmatic nucleus (SCN), the master pacemaker in the hypothalamus that receives light input from the retina and synchronizes circadian oscillators in other brain regions and peripheral tissues. Lacking the tight neuronal network that couples single‐cell oscillators in the SCN, circadian clocks outside the SCN may be less stable and more susceptible to disturbances, for example by clock gene mutations or uncontrollable stress. However, non‐SCN circadian clocks have not been studied extensively in rodent models of mood disorders. In the present study, it was hypothesized that disturbances of local circadian clocks in mood‐regulating brain areas are associated with depression‐like behaviour in mice. Using the learned helplessness procedure, depression‐like behaviour was evoked in mice bearing the PER2::LUC circadian reporter, and then circadian rhythms of PER2 expression were examined in brain slices from these mice using luminometry and bioluminescence imaging. It was found that helplessness is associated with absence of circadian rhythms in the nucleus accumbens and the periaqueductal grey, two of the most critical brain regions within the reward circuit. The current study provides evidence that susceptibility of mice to depression‐like behaviour is associated with disturbed local circadian clocks in a subset of mood‐regulating brain areas, but the direction of causality remains to be determined.     

Oligomeric proanthocyanidin protects retinal ganglion cells against oxidative stress-induced apoptosis

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药用植物组织培养中褐化现象的研究进展

药用植物外植体组织培养阶段,易受到各种因素的影响而产生褐化现象,这不仅影响药用植物外植体的鲜活度,而且还干扰了愈伤组织的诱导和再生,严重阻碍了药用有效成分的快速积累和组培技术的成功率。因此,研发快速、有效、安全、经济的褐化抑制方法显得尤为重要。对药用植物褐化现象、褐化机制、褐化影响因素及防褐有效措施进行了综述,以期为药用植物组织培养研究中抑褐措施提供科学参考,并对药用植物组织培养防褐技术进行展望。     

Milk Thistle Extract and Silymarin Inhibit Lipopolysaccharide Induced Lamellar Separation of Hoof Explants in Vitro

The pathogenesis of laminitis is not completely identified and the role of endotoxins (lipopolysaccharides, LPS) in this process remains unclear. Phytogenic substances, like milk thistle (MT) and silymarin, are known for their anti-inflammatory and antioxidant properties and might therefore have the potential to counteract endotoxin induced effects on the hoof lamellar tissue. The aim of our study was to investigate the influence of endotoxins on lamellar tissue integrity and to test if MT and silymarin are capable of inhibiting LPS-induced effects in an in vitro/ex vivo model. In preliminary tests, LPS neutralization efficiency of these phytogenics was determined in an in vitro neutralization assay. Furthermore, tissue explants gained from hooves of slaughter horses were tested for lamellar separation after incubation with different concentrations of LPS. By combined incubation of explants with LPS and either Polymyxin B (PMB; positive control), MT or silymarin, the influence of these substances on LPS-induced effects was assessed. In the in vitro neutralization assay, MT and silymarin reduced LPS concentrations by 64% and 75%, respectively, in comparison PMB reduced 98% of the LPS concentration. In hoof explants, LPS led to a concentration dependent separation. Accordantly, separation force was significantly decreased by 10 µg/mL LPS. PMB, MT and silymarin could significantly improve tissue integrity of explants incubated with 10 µg/mL LPS. This study showed that LPS had a negative influence on the structure of hoof explants in vitro. MT and silymarin reduced endotoxin activity and inhibited LPS-induced effects on the lamellar tissue. Hence, MT and silymarin might be used to support the prevention of laminitis and should be further evaluated for this application.     

Growth of pyramidal, but not non‐pyramidal, dendrites in long‐term organotypic explants of neonatal rat neocortex chronically exposed to neurotrophin‐3

The present study was undertaken to determine the effects of neurotrophin-3 (NT3) and spontaneous bioelectric activity (SBA) on dendritic elongation and branching in long-term isolated organotypic explants of rat neocortex. Viral vector-directed expression of NT3 was used as an effective means to ensure a continuous, local production of the neurotrophic factor. Quantitative light microscopic measurement of dendritic branching patterns was carried out on Golgi-stained materials. Explants were exposed to an adenoviral vector encoding the genetic sequence for neurotrophin-3 (Ad-NT3), or to exogenous additions of the neuropeptide NT3. In order to test for activity-dependent growth effects under control and experimental conditions, explants were exposed to glutamatergic blockade using a cocktail of APV and DNQX. Both Ad-NT3 and NT3 peptide potently promoted apical and basal dendritic growth (elongation and branching) in pyramidal neurons. This growth was observed to be significant in layers II–IV and V. These growth effects were also not activity dependent, inasmuch as they were elicited from explants in which spontaneous bioelectric activity had been suppressed. Non-pyramidal neurons, throughout the neocortical slice, showed no significant dendritic responses to the prolonged presence of NT3. These findings show that pyramidal dendritic growth in long-term neocortical explants responds to at least one neurotrophic growth factor, NT3, and is independent of intrinsic bioelectric activity. The use of viral vectors in delivering a continuous high level of neurotrophic factor within developing neural tissues demonstrates its potential application to in vivo tissues during development, or in the stimulation of neuritogenesis and neuroregeneration following injuries.     

Biosynthesis and partial characterization of tear film glycoproteins. Incorporation of radioactive precursors by human lacrimal gland explants in vitro

A system for studying the biosynthesis of tear glycoproteins by human lacrimal gland is described. Sufficient quantities of the newly-synthesized glycoproteins were obtained to permit some analyses of their physical and chemical properties. Separation of glycoproteins by gel filtration on Sephadex G-200 showed three major heterogeneous glycoprotein fractions with molecular weights of 580 000, 100 000 and 39 000 daltons. The sulfate, carbohydrate, and amino acid composition of each fraction was determined. The protein moieties were found to be acidic in nature. The smaller molecular weight fractions were moderately richer in carbohydrate [26% (w/w) than the higher molecular weight fractions (8% w/w)]. The low carbohydrate contents were comparable to those of plasma-type proteins. DEAE ion exchange chromatography of the major fraction also showed an elution pattern characteristic of plasma-type proteins. Using immunodiffusion techniques, at least five serum-type proteins, including IgA, IgG, albumin, transferrin and cerulo-plasmin were identified from the different fractions of the cultured lacrimal gland secretions. It appeared that human lacrimal gland secreted similar plasma proteins to those found in the tears. The significance of these identified tear proteins to the physiology of tears is discussed.     

Acid phosphatase activity: a marker of androgen action in prostate explant cultures

Acid phosphatase activity in rat ventral prostate explants has been assayed to determine if this parameter could serve as a specific and quantitative marker of androgen action in this in vitro model. Dihydrotestosterone (10 nM) caused an absolute increase in both total (42.5 +/- 2.9 vs control 27.1 +/- 4.0 nmoles p-nitrophenol generated in 30 min/micrograms DNA, P less than .01) and tartrate-resistant acid phosphatase activity (34.1 +/- 1.5 vs control 17.2 +/- 2.8 U/micrograms DNA, P less than .05), and this effect was maximal on the 4th day of culture. This was the time when explant weight and DNA content tended to fall or only to be maintained by androgen. Similar changes were observed with the potent synthetic androgen, mibolerone. The addition of either the antiandrogen cyproterone acetate or flutamide in a 100-fold excess to that of androgen caused significant inhibition in acid phosphatase activity. No significant change was observed at low concentrations of estradiol or progesterone, and only minimal and inconsistent increases in activity were noted at high concentrations. No increase was noted when cortisol, cyproterone acetate, or flutamide was added to the media. We conclude that measurement of acid phosphatase activity in cultured explants of rat ventral prostate provides a biochemical marker of androgenicity that is more specific than measurement of [3H]-thymidine incorporation.