Gene delivery to skeletal muscle is a promising strategy for the treatment of muscle disorders and for the systemic secretion of therapeutic proteins. However, present DNA delivery technologies have to be improved with regard to both the level of expression and interindividual variability. We report very efficient plasmid DNA transfer in muscle fibers by using square-wave electric pulses of low field strength (less than 300 V͞cm) and of long duration (more than 1 ms). Contrary to the electropermeabilization-induced uptake of small molecules into muscle fibers, plasmid DNA has to be present in the tissue during the electric pulses, suggesting a direct effect of the electric field on DNA during electrotransfer. This i.m. electrotransfer method increases reporter and therapeutic gene expression by several orders of magnitude in various muscles in mouse, rat, rabbit, and monkey. Moreover, i.m. electrotransfer strongly decreases variability. Stability of expression was observed for at least 9 months. With a pCMV-FGF1 plasmid coding for fibroblast growth factor 1, this protein was immunodetected in the majority of muscle fibers subjected to the electric pulses. DNA electrotransfer in muscle may have broad applications in gene therapy and in physiological, pharmacological, and developmental studies.

Polyclonal antibodies exclusively recognizing A beta 40 (FCA 3340) or A beta 42 (FCA3542) were obtained. These demonstrated that FAD-linked presenilins similarly affect both p342 and A beta 42, suggesting that these mutations misroute the beta APP to a compartment where gamma-secretase, but not alpha-secretase, cleavages are modified. Overall, these antibodies should prove useful for fundamental and diagnostic approaches, as suggested by their usefulness for biochemical, cell biological, and immunohistochemical techniques.

Numerous diseases are linked to the absence or insufficient concentration of a specific plasma protein. Gene transfer is an appealing strategy for correction of such diseases. We report high and sustained plasma secretion of human secreted alkaline phosphatase and of human Factor IX by skeletal muscle of mice. This was obtained by delivering square-wave unipolar electric pulses of low field strength (200 V/cm) and long duration (20 ms) to skeletal muscle previously injected with plasmid DNA encoding for the secreted protein. This intramuscular electrotransfer method allows 30- to 150-fold increase in reporter protein secretion, compared to simple plasmid DNA injection. This increase allows one to obtain values of up to 2200 ng/ml of a reporter circulating protein. Moreover, this high level of secretion remains stable for several months.

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Minicircles are a new form of supercoiled DNA molecule for enhancer/promoter. Comparing maximal differences, these nonviral gene transfer which have neither bacterial origin of minicircles gave 2.5 to 5.5 times more reporter gene replication nor antibiotic resistance marker. They are thus activity than the unrecombined plasmid in the NIH3T3 cell smaller and potentially safer than the standard plasmids line and rabbit smooth muscle cells. Moreover, injection in currently used in gene therapy. They were obtained in E.vivo into mouse cranial tibial muscle, or human head and coli by att site-specific recombination mediated by the neck carcinoma grafted in nude mice resulted in 13 to 50 phage integrase, which was used to excise the times more reporter gene expression with minicircles than expression cassette from the unwanted plasmid with the unrecombined plasmid or larger plasmids. Histosequences. We produced two minicircles containing the logical analysis in muscle showed there were more transluciferase or ␤-galactosidase gene under the control of fected myofibers with minicircles than with unrecombined the strong human cytomegalovirus immediate-early plasmid.

beta A4 immunoreactivity was studied in temporal neocortex, area 22, of 26 cases with graded intellectual status. Sampling was performed in psychometrically assessed women over 75 years, either intellectually normal or affected by senile dementia of Alzheimer type of various degrees of severity. beta A4 antibodies labelled various types of beta A4 deposits in 22/26 cases: (1) small, stellate deposits; (2) diffuse deposits, (3) primitive, (4) classic and (5) compact, or burn-out, plaques. The densities of the stellate deposits, primitive and classic plaques were always positively linked with the severity of the intellectual status, whereas those of the diffuse deposits were not. This was due to a single case with normal mental status and numerous beta A4 deposits. Densities of stellate and diffuse deposits were higher in layers I, III and IV, whereas densities of primitive, classic, and neuritic plaques observed with Bodian's technique were higher in layers II and III. Topographical distribution of each subtype did not vary as a function of the severity of the intellectual status. These data suggest that deposits of beta A4 protein appear a necessary but not a sufficient condition for inducing neuritic plaque formation, in the neocortex as in other brain areas. beta A4 proteins could accumulate either as diffuse deposits, which do not cause an intellectual deficit, or as dense deposits, associated with argyrophilic neurites, i.e., classic neuritic plaques, highly correlated to the intellectual impairment. This evolution could depend on factors which are laminarily distributed in the neocortex.

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