We have isolated and sequenced a full-length cDNA clone encoding rat glyceraldehyde-3-phosphate-dehydrogenase (GAPDH, E.C.1.2.1.12). The entire mRNA is 1269 nucleotides long exclusive of poly(A) and contains respectively 71 and 196 bases of 5' and 3' non-coding regions. Primer extension as well as S1 nuclease protection experiments clearly established that a single (or at least a highly prominent) GAPDH mRNA species is expressed in all rat tissues examined. This sequence allowed the determination of the hitherto unknown primary structure of rat GAPDH which is 333 aminoacids long. Comparison between GAPDH sequences from rat, man and chicken revealed a high degree of sequence conservation at both nucleotide and protein levels.

Here, we report the isolation of a human multipotent adipose-derived stem (hMADS) cell population from adipose tissue of young donors. hMADS cells display normal karyotype; have active telomerase; proliferate >200 population doublings; and differentiate into adipocytes, osteoblasts, and myoblasts. Flow cytometry analysis indicates that hMADS cells are CD44+, CD49b+, CD105+, CD90+, CD13+, Stro-1−, CD34−, CD15−, CD117−, Flk-1−, gly-A−, CD133−, HLA-DR−, and HLA-Ilow. Transplantation of hMADS cells into the mdx mouse, an animal model of Duchenne muscular dystrophy, results in substantial expression of human dystrophin in the injected tibialis anterior and the adjacent gastrocnemius muscle. Long-term engraftment of hMADS cells takes place in nonimmunocompromised animals. Based on the small amounts of an easily available tissue source, their strong capacity for expansion ex vivo, their multipotent differentiation, and their immune-privileged behavior, our results suggest that hMADS cells will be an important tool for muscle cell–mediated therapy.

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The The lipostasis theory postulates that the size of the body fat depot is regulated by a feedback loop (1). Body weight is lightly regulated in vivo, and the original fat cell mass is precisely reconstituted after lipectomy in adults (2). These findings suggest that the feedback loop operates at a set level in each individual.The recently cloned mouse obese gene (ob) appears to encode a fat cell signal in this feedback loop (3). Adipocytes transplanted from genetically obese C57BJ/6J ob/ob mice into wild-type animals (or from wild-type mice into ob/ob mice) ultimately achieve the same lipid content as adipocytes from the recipient animal (4,5). This result suggests that the function of ob is not autonomous to fat cells.If the ob gene encodes a signal that acts at a distant site to regulate the overall size of the body's lipid stores, (i) this signal should be made in adipocytes, the principal site of lipid storage, (ii) this signal should be made in all adipose tissue depots, and (iii) a secondary increase in ob expression level should be associated with defects downstream of ob in the pathway(s) that control adiposity. In this paper, we present data consonant with these predictions. MATERIALS AND METHODSIn Situ Hybridization. White fat tissues from identical abdominal regions of wild-type (wt) and db mice were processed simultaneously according to the modified method de (Fig. 1, labeled wt). Signals were not noted when the antisense probe was hybridized to brain sections (data not shown). Hybridization of the antisense probe to sections of adipose tissue from C57BL/Ks db/db mice was greatly increased, confirming the adipocyte-specific expression of ob RNA and demonstrating a large increase in the level of ob RNA per adipocyte in these animals ( Fig. 1, labeled db/db). Mice mutant at the db locus are massively obese as part of a syndrome that is phenotypically identical to that seen in C57BL/6J ob/ob mice (12).ob RNA was not synthesized by adipose tissue stromal cells separated from adipocytes. As expected, cells in the adipocyte fraction expressed ob RNA using Northern blots (Fig. 2). The same result was obtained using reverse-transcription-PCR (data not shown). These data support the conclusion that only adipocytes express the ob gene. Data from cultured adipocytes Abbreviations: GTG, gold thioglucose; wt, wild type.§To whom reprint requests should be sent at the t address.6957

To address the possibility that the expression of the myc gene might be regulated at a post-transcriptional level, we have investigated the half-life of myc mRNA in various cells. Our survey included normal human embryonic fibroblasts as well as transformed human cells of various origins: cervix carcinoma (HeLa), breast carcinoma (MCF7), Burkitt lymphoma (Daudi), and promyelocytic leukemia (HL60). All these cells revealed an extreme instability of myc mRNA (half-life, 10 min), suggesting that the control of myc mRNA degradation might be a general means (although not necessarily exclusive) of regulating both the level and the timing of myc gene expression. Inhibition of protein synthesis resulted in a dramatic stabilization of myc mRNA in HeLa, MCF7, and HL60 cells, suggesting that the controlling element might itself be, at least in these cells, a protein of rapid turnover. This finding opens the way to studying the mechanism of myc mRNA inactivation in these different cell types. However, protein synthesis inhibition had no effect on myc mRNA instability in other transformed (Daudi) cell lines as well as normal embryonic human fibroblasts. These different types of behavior suggest that the post-transcriptional control of myc gene expression might involve multiple factors that would be differently affected in various cell types.

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