e adult moth were collected, and frozen immediately in liquid nitrogen. Total mRNA was extracted using Trizol reagent, and DNA contamination in the mRNA sam ples was digested with RNase free DNase I. The concentration of RNA was calculated by spectrophotometry. The first strand of cDNA was synthesized from 1 ug mRNA using M MLV Reverse Transcriptase following the man ufacturers instructions. Verification of the putative silkworm apoptosis related genes The PCR primers were designed based on the coding sequences of the putative silkworm apoptosis related genes identified by the bio informatics analysis. performed in a total reaction volume of 25 ul, containing normalized cDNA, 15 pmol of each primer, 2 mM MgCl2, 0. 25 mM dNTP, 1�� buffer, 2. 5 units of Taq DNA polymerase and distilled deionized H2O.

PCR was performed as follows, initial denaturation at 94 C for 3 min, followed by 25 cycles of 30 s each at 94 C, 1 min annealing, 1 3 min extension at 72 C, and a final extension at 72 C for 10 min. The amplification products Batimastat were analyzed on 1% agarose gels, and sequenced and confirmed by the Ying Jun Company and Bio engineering. Analysis of apoptosis related genes of different developmental stages More than 184201 ESTs from Bombyx mori are available in the NCBI database. To search transcripts for individual apoptosis related genes, a BlastN search was conducted against the silkworm EST database. The putative coding sequences were used as queries. A 95% or greater identity and minimum cut off E value were employed to discriminate between duplicated genes.

Microarray data analysis was performed as described by Xia and colleagues. Androgens have been shown to reverse muscle loss due to age, and to preserve muscle in persons with HIV infection and burns. In animal models, androgens also prevent or reduce atrophy due to disuse from spinal cord injury, immobilization, or unweighting. The molecular basis for these beneficial effects remains poorly understood. One major factor that contributes to muscle atrophy is accelerated catabolism of muscle proteins, which is lar gely attributable to the ubiquitin proteasome pathway and which has been linked to the muscle ubiquitin E3 ligases muscle atrophy F box and muscle Ring finger 1. Upregulation of MAFbx and MuRF1 has been attributed to activation of FOXO1.

Degradation by MAFbx of the muscle differen tiation factor MyoD or the translation initiation fac tor eIF3F have also been linked to muscle atrophy. In cardiac myocytes, MAFbx also reduces calcium dependent signaling through calcineurin and has been shown to reduce myocyte size. A role has been established in muscle atrophy for inhibi tors of protein synthesis acting both up and downstream of mTOR, a protein kinase that integrates signals regulat ing protein synthesis and cell size and has also been impli cated in muscle hypertrophy. Reductions in mTOR activity caused by dexamethasone or ethanol have been shown to be due to upregulation of REDD1. mTOR is also