With the goal of performing astrocyte-specific modification of genes in the mouse, we have generated a transgenic line expressing Cre recombinase under the control of the human glial fibrillary acidic protein (hGFAP) promoter. Activity was monitored by crossing the hGFAP-cre transgenics with either of two reporter lines carrying a lacZ gene whose expression requires excision of loxP-flanked stop sequences. We found that lacZ expression was primarily limited to the central nervous system, but therein was widespread in neurons and ependyma. Cell types within the brain that notably failed to activate lacZ expression included Purkinje neurons of the cerebellum and choroid plexus epithelium. Onset of Cre expression began in the forebrain by e13.5, suggesting that the hGFAP promoter is active in a multi-potential neural stem cell.
Using a human glial fibrillary acidic protein (hGFAP) promoter-driven cre transgene, we have achieved efficient inactivation of a floxed connexin43 (Cx43) gene in astrocytes of adult mice. The loss of Cx43 expression was monitored in a cell-autonomous manner via conditional replacement of the Cx43-coding region by a lacZ reporter gene. In this way, we bypassed the early postnatal lethality previously reported for Cx43 null mice and characterized the phenotypic consequences of Cx43 deficiency in the CNS. Mice lacking Cx43 in astrocytes were viable and showed no evidence of either neurodegeneration or astrogliosis. Spreading depression (SD) is a pathophysiological phenomenon observed in the CNS that is characterized by a propagating wave of depolarization followed by neuronal inactivation. Inhibitors of gap junctional communication have previously been shown to block initiation and propagation of SD. In contrast, we observed an increase in the velocity of hippocampal SD in the stratum radiatum of mice lacking Cx43 in astrocytes. In the same brain subregion, dye-coupling experiments revealed a reduction in overall astrocytic intercellular communication by approximately 50%. This strongly suggests separate and different neuronal and glial contributions of gap junctional intercellular communication to SD. Concomitant with increased velocity of spreading depression, we observed enhanced locomotory activity in mice lacking Cx43 in astrocytes.
Green fluorescent protein (hGFP-S65T) was expressed in transgenic mice under the control of the astrocyte-specific glial fibrillary acidic protein (GFAP) promoter. Tissues from two independent transgenic lines were characterized by Northern blot analysis and by confocal microscopy. The expression pattern in these two lines was identical in all tissues examined, and similar to that found previously with a lacZ transgene driven by the same promoter. Bright fluorescence was observed in the cell bodies and processes of unfixed or fixed astrocytes, using both whole mount and brain slice preparations, from multiple areas of the central nervous system. However, in contrast to GFAP-lacZ transgenics, retinal Müller cells expressed the GFP transgene in response to degeneration of neighboring photoreceptors. These data indicate that the 2.2-kb hGFAP promoter contains sufficient regulatory elements to direct expression in Müller cells, and that GFP is a suitable reporter gene for use in living preparations of the mammalian nervous system. Such mice should prove useful for studies of dynamic changes in astrocyte morphology during development, and in response to physiological and pathological conditions.
Aim: Thalassaemia is a good candidate disease for control by preventive genetic programmes in developing countries. Accurate population frequency data are needed for planning the control of thalassaemia in the high risk Guangdong Province of southern China. Methods: In total, 13 397 consecutive samples from five geographical areas of Guangdong Province were analysed for both haematological and molecular parameters.Results: There was a high prevalence of carriers of a thalassaemia (8.53%), b thalassaemia (2.54%), and both a and b thalassaemia (0.26%). Overall, 11.07% of the population in this area were heterozygous carriers of a and b thalassaemia. The mutation spectrum of a and b thalassaemia and its constitution were fully described in this area. This study reports the true prevalence of silent a thalassaemia in the southern China population for the first time. In addition, two novel mutations that give rise to a thalassaemia, one deletion resulting in b thalassaemia, and a rare deletion (22 THAI allele) previously unreported in mainland China were detected. The frequency of the most common mutation, the Southeast Asian type of deletion (22 SEA , accounting for 48.54% of all a thalassaemias) was similar to the total of two a + thalassaemia deletions (2a 3.7 and 2a 4.2 , accounting for 47.49% of a thalassaemia). Conclusion: Both a and b thalassaemia are widely distributed in Guangdong Province of China. The knowledge gained in this study will enable the projected number of pregnancies at risk to be estimated and a screening strategy for control of thalassaemia to be designed in this area.
In designing drug carriers, the drug-to-carrier ratio is an important consideration because using high quantities of carriers can cause toxicity resulting from poor metabolism and elimination of the carriers1. However, these issues would be of less concern if both the drug and carrier possess therapeutic effects. (-)-Epigallocatechin-3-O-gallate (EGCG), which is a major ingredient of green tea, has been shown to possess anticancer effects2-7, anti-HIV effects8, neuroprotective effects9, DNA-protective effects10, etc. Here we show that sequential self-assembly of the EGCG derivative with anticancer proteins forms stable micellar nanocomplexes (MNCs), which have greater anticancer effects in vitro and in vivo than the free protein. The MNC is obtained by complexation of oligomerized EGCG with the anticancer protein, Herceptin, to form the core, followed by complexation of poly(ethylene glycol)-EGCG to form the shell. When injected into mice, the Herceptin-loaded MNC showed better tumour selectivity and growth reduction, and longer blood-half-life than free Herceptin.
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