Accumulation Of Acidic Protein Research Articles

Distribution of glial fibrillary acidic protein accumulation after retinal tack insertion for intraocular fixation of epiretinal implants

surface of the globe. This dissection was similarly continued until past the level of the fornices. The fatty tissue and Tenon’s capsule behind the conjunctiva were then divided until the anterior and posterior planes met, as demonstrated in Fig. 1I. It was found that it was easier to perform this dissection from the anterior plane using scissors to cut down onto the globe, taking care to do so beyond the fornices, so as not to breach the conjunctiva. This was completed circumferentially allowing the conjunctiva to be excised as a whole specimen. Eye sockets were reconstructed using cotton wool and plastic eye shields that hold the eyelids in a closed position in accordance with the guidance standards of the Royal College of Ophthalmologists (2008) with good aesthetic results. Using this technique, whole conjunctival tissue was successfully obtained from 16 human cadaveric donors (32 eyes). Conjunctival tissue was subsequently wax embedded, sectioned and examined histologically confirming the presence of tarsal to limbal conjunctiva (Fig. 1J). Conjunctival epithelial cells were also harvested using trypsin disaggregation and co-cultured on an inactivated J23T3 mouse fibroblast feeder layer. Cultured cells were fixed in methanol and immunofluorescently stained for the conjunctival marker cytokeratin 19 (R. M. K. Stewart unpublished data). We have developed a technique to retrieve whole human cadaveric conjunctiva. This will further enable future ex vivo and in vitro analyses of the whole human conjunctiva, which require a continuum of conjunctival tissue. This is of particular benefit to studies assessing the distribution and characterization of conjunctival stem cells to aid propagation and transplantation; similarly for inflammatory cells and vasculature to improve knowledge of ocular surface immunity and potential for therapeutic agents.

Induction of heat-shock Balbiani Rings after RNA synthesis inhibition in polytene chromosomes of Chironomus thummi

We have studied the effect of RNA synthesis inhibitors, such as Actinomycin D, α-Amanitin, or DRB, on the heat-inducible puffs of Chironomus thummi. While the appearance of most of the T-puffs is blocked by the inhibitory treatments, the induction of a heat-shock Balbiani Ring (T-BR-III) takes place in the presence of these drugs. No transcriptional activity could be detected in this structure. However, its frequency and approximate size values are similar to those obtained for T-BR-IIIs induced under normal conditions. Accumulation of acidic proteins also takes place in T-BR-IIIs in the presence of these inhibitors. On the other hand, puff III-A3b, a major T-puff of Ch. thummi, seems to be resistent to DRB treatment at both the puffing and the transcriptional levels. These results are discussed in the context of puffing-RNA synthesis relationships.

The synthesis of acidic chromosomal proteins during the cell cycle of HeLa S-3 cells. I. The accelerated accumulation of acidic residual nuclear protein before the initiation of DNA replication.

The synthesis and accumulation of acidic proteins in the tightly bound residual nuclear fraction goes on throughout the cell cycle of continuously dividing populations of HeLa S-3 cells; however, during late G(1) there is an increased rate of synthesis and accumulation of these proteins which precedes the onset of DNA synthesis. Unlike that of the histones, whose synthesis is tightly coupled to DNA replication, the synthesis of acidic residual nuclear proteins is insensitive to inhibitors of DNA synthesis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of acidic residual nuclear proteins shows different profiles during the G(1), S, and G(2) phases of the cell cycle. These results suggest that, in contrast to histones whose synthesis appears to be highly regulated, the acidic residual proteins may have a regulatory function in the control of cell proliferation in continuously dividing mammalian cells.

Factors involved in the expression of gene activity in polytene chromosomes.

In order to separate some of the factors involved in the formation of puffs the antibiotic actinomycin D was applied at different stages of puff activity. Puffs were induced by temperature shocks or eodysone. Inhibition of RNA synthesis with actinomycin D before application of a puff inducing stimulus prevents neither the appearance of the stimulus specific puffs nor the accumulation of acidic proteins in the puff regions. The puffs attained under these conditions approximately 1/3 of the size normally produced by the stimulus. Indications were obtained that during puff formation acidic protein accumulation precedes the onset of RNA synthesis. Synthesis and storage of newly synthesized RNA within the puff region was studied on the basis of grain distribution in uridine-H3 autoradiographs after various incubation periods. RNA synthesis appears to be restricted to a particular area of the puff region. After a 3 min temperature shock following injection of uridine-H3 silver grains are located only over a particular area of the newly formed puff. The same area becomes labeled during a 1 min pulse of uridine-H3 applied at a stage of maximum puff development. Longer periods of incubation result in a random distribution of the grains over the whole puff region. Grain counts on different areas of experimentally induced puffs and on the same areas at a stage of puff regression indicate that the newly synthesized RNA becomes transferred from the area where it was synthesized and is stored for a certain period within the puff region. Complete release of newly synthesized RNA from puffs in which RNA synthesis was inhibited by actinomycin D at a stage of maximal activity is accomplished within 30 to 35 min.

The hormone ecdysone as effector of specific changes in the pattern of gene activities of Drosophila hydei.

The hormone ecdysone induces a large number of changes in the puffing pattern of mid third instar larvae of Drosophila hydei. The pattern of changes occurring after experimental administration of the hormone are identical with those observed in normal development during a 6 hour period before puparium formation. After administration of the hormone a considerable number of puffs react with a change in activity within 15–20 min. During this period 3 puffs arise newly, 12 puffs show a strong increase in activity, 6 puffs show a less pronounced increase in activity and 12 puffs show a decrease in activity. At a period of 4–6 hours after administration of the hormone another 5 puffs arise newly. The effect of the hormone was identical in both in vivo and in vitro experiments. — Diameter measurements on several puffs reacting within 30 min with an increase in diameter showed that these puffs reacted simultaneously. Most of the puffs that showed a decrease in activity reacted with some delay. — A study of the effect of different hormone concentrations revealed that the kinetics of 4 puffs with respect to the relationship between concentration and puff size was identical over a range of concentrations from 33·10−5 to 33CU/μl. Three of these puffs showed a reaction with even lower concentrations. Maximum puff size is attained by all puffs at a concentration of 33·10−4CU/μl. Among the puffs studied no difference in their reaction threshold was found. — A study of the behavior of 5 puffs of the group reacting within 15–20 min and one of the group reacting after 4–6 hours in midintestine and Malpighian tubules revealed that these puffs showed the same reaction after injection of the hormone as observed in the salivary glands. — All puffs activated by administration of the hormone showed particularly strong uptake of tritiated uridine and accumulation of acidic protein. — It is concluded that the hormone ecdysone induces a pattern of changes in gene activity that is far more complex in Drosophila hydei than in Chironomus tentans.