实验仪器使用技术

Many of the early studies on the localization of calpain described a cytosolic distribution. However, it is becoming clear that calpain can play an essential role in regulating signal transduction across members of the integrin family of adhesion receptors and that in order to do this it is recru ...

There is now a considerable literature suggesting that the calpain/calpastatin system may be involved in myogenesis (1–7). Primary myoblasts in culture at first proliferate, but then usually cease to divide, in response to complex signals. The cells then migrate, align, and fuse to form the mu ...

A major area of calpain research involves the development of techniques to assess calpain activation in cells. Knowing physiologic conditions that allow the various calpain isoforms in cells to become activated should provide valuable information about their roles in cell functio ...

Cell membrane fusion is a ubiquitous cellular process, mediating such phenomena as fertilization, muscle development, certain viral infections, and giant cell formation from macrophages. For membrane fusion to occur, the cell membranes must attach to each other, usually by means of in ...

Calpain is a calcium-activated neutral proteinase present in all mammalian tissues thus far studied (1,2). Ubiquitous calpain exists as μ-calpain and m-calpain isoforms which require μM and mM calcium levels for activation, respectively. In the central nervous system (CNS) μ-calpain is ...

Identification of the preferred brain substrates of calpain in vivo is not an easy task. The first difficulty is that, given enough time, almost any protein will be degraded when incubated in the presence of calpain and Ca+ in a test tube. A second major problem is the lack of completely specific calpain in ...

The calpains are found ubiquitously in mammalian cells and are activated following various central nervous system (CNS) insults, including ischemia (1), spinal cord injury (2) and traumatic brain injury (TBI) (3–5). Nonerythroid α-spectrin is a submembrane cytoskeletal protein and p ...

The preparation of freshly isolated rat renal proximal tubules in suspension has been utilized by our laboratory for more than 10 yr and reported in numerous publications (1–10). This method of tubule isolation by collagenase digestion and Percoll centrifugation has recently been used by ...

The general assay for proteolytic activity using casein as a substrate was described in 1947. A modification using azocasein has been described, in which digestion by calpain results in colored peptides soluble in trichloroacetic acid (TCA), so that the intensity of the color in the supernat ...

Although considerable information about calpain has been obtained from studies using isolated enzyme preparations, this does not permit the dynamic assessment of intracellular protease activity during changes in intracellular Ca2+ and pH. Intracellular calpain activity is ...

Ischemia-reperfusion (I/Rp) injury to the liver occurs after liver transplantation, shock or in surgical procedures in which vascular supply to the liver is temporarily abrogated. The extent of I/Rp-mediated injury is dependent on several factors including duration of ischemia, and in ...

Calpain proteases have been extensively studied over the past 20 years but substrate specificity and activity studies have been limited to investigations using isolated calpain proteases in vitro and in the presence of calcium concentrations beyond the physiologic range found in no ...

Erythrocyte spectrin and nonerythroid spectrin (fodrin, calspectin) are major constituents of the membrane cytoskeleton, forming a two-dimensional meshwork beneath the outer cell membrane (1,2). The three-dimensional organization of the membrane cytoskeleton has been ex ...

Many types of cataracts in the lens of the eye show elevated concentrations of calcium, which could activate calpains, and thus this chapter presents the techniques specialized for measuring calpain activity in lens. Ubiquitous calpain has been assayed in lenses from man, cow, pig, rat, sheep, m ...

Circulating platelets undergo dramatic morphological changes in response to platelet agonists, resulting in attachment, spreading, secretion, and aggregation. These rapid dynamic physiological activities require a series of rapid biochemical reactions, including ty ...

An inherent consequence of calpain activation in situ is activation of the ubiquitous signal transduction kinase, protein kinase C (PKC). Whether or not one is directly interested in the effects of PKC, it may be important during experimental design and interpretation to consider that cert ...

p53 is a key regulator of the cell cycle and apoptosis which is essential for maintenance of genome integrity (1–4). It can function as a transcription factor, but other roles in DNA repair, homologous recombination and the regulation of its own mRNA translation have been proposed. p53 has also been re ...

The goals of our research program have been to understand the mecha-nism(s) responsible for the turnover of myofibrillar proteins of skeletal muscle. This understanding would be useful in development of strategies to treat muscle wasting diseases and, possibly, to augment efficiency of ...

The phenomenon of apoptosis has gone from obscurity in 1972, when it was named (1), to the limelight of over 6000 publications in 1998 alone. Originally described as a peculiar morphology of cell death, seen when that death was “physiological” or programmed, apoptosis remains most clearly deter ...

Neurotrophins are a family of growth factor proteins sharing a considerable degree of primary sequence and tertiary structure homology (1–4). Members of this protein family, including nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and ...