Physiological cell death during development and its relationship to aging.
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Aging↗
Biomedical subjects
Z Zakeri
Publications and source records attributed to Z Zakeri.
Apoptotic cell death in the mouse limb and its suppression in the hammertoe mutant.
We examined the importance of cell death in producing the limb deformity phenotype of the Hammertoe mouse. In order to make this evaluation, we first combined several techniques to verify the reliability of cell death markers in the normal mouse limb. In the normal limb, cell death occurs between Days 10.5 and 15.5 in the anterior, interdigital, and posterior zones. Morphological examination and staining for acid phosphatase or macrophages all reveal regions of cell death by detecting phagocytosis, while in situ end labeling of DNA defines nuclear degeneration as apoptotic. The phagocytes display surface markers characteristic of macrophages. The Hammertoe mutant shows strongly reduced cell death only in the interdigital region. This result is consistent with the phenotype and suggests that the mutation influences the patterning or control of cell death rather than its mechanism.
Animals↗
Rat Sertoli and spermatogenic cells express a similar gene, and its product is antigenically related to an outer dense fiber-associated protein.
We have previously reported that a heterodimeric protein secreted by rat Sertoli cells is antigenically related to a protein associated with outer dense fibers of the sperm tail. Therefore, we have explored the possibility that Sertoli and spermatogenic cells express a similar gene encoding a homologous protein. A Sertoli cell heterodimeric protein cDNA probe recognizes specific mRNA in pachytene and round spermatids fractionated by centrifugal elutriation; however, this specific mRNA was less prominent than in cultured Sertoli cells. In agreement with these observations, in situ hybridization experiments show that Sertoli cells are predominantly engaged in active heterodimeric protein mRNA synthesis, while meiotic prophase spermatocytes and spermatids also show significant but less abundant specific mRNA. Immunoblotting experiments demonstrate that, while Sertoli cells synthesize a heterodimeric protein consisting of two disulfide-linked components with molecular masses of 45 and 35 kD, both primary spermatocytes and round spermatids synthesize single 30 kD monomers not associated by disulfide linkage but recognized by antisera to Sertoli cell heterodimeric protein. Immunoblotting and immunogold electron microscopic studies show that antisera to Sertoli cell heterodimeric protein recognize a protein associated with outer dense fibers. This immunoreactivity was abolished by a 5-min pronase treatment, without affecting the integrity of outer dense fibers. Results of this study and previous studies demonstrate that both Sertoli and spermatogenic cells express a similar gene and that an antigenically related product encoded by this gene becomes associated with outer dense fibers during their assembly at spermiogenesis.
Animals↗
Developmental expression of the S35-S45/SGP-2/TRPM-2 gene in rat testis and epididymis.
Testosterone-repressed prostate message-2 (TRPM-2) was originally isolated and cloned from the regressing ventral prostate of the rat. In this tissue, and in other hormone-dependent tissues such as the mammary gland, this gene is induced in the absence of the appropriate trophic hormone. Sequence analysis of the cDNA and genomic clones of TRPM-2 have demonstrated that the coding sequence of this gene is identical to S35-S45 (also known as SGP-2 and clusterin), which is constitutively expressed by the Sertoli cells of the adult testis. Using Northern, slot blot, S1-nuclease analysis, and in situ hybridization, we have investigated the regulation of TRPM-2 expression in the testis and epididymis during development. Slot blot analysis of RNA extracted from the testis and epididymis of 7-, 14-, 28-, 35-, and 91-day-old rats demonstrates that the gene is induced to detectable levels between days 7 and 14 and that the relative level of expression does not change significantly after day 14. In situ hybridization using frozen sections of testis from day 2-, 7-, 14-, 28-, 35-, and 91-day-old rats confirms that there is little expression of TPRM-2 in the seminiferous epithelium of 7-day-old rats, but this increases considerably after 14 days, primarily in Sertoli cells but also in association with meiotic developing spermatogenic cells. However, TRPM-2 mRNA is expressed in the rete testis at 2 days of age, reaches a peak at 35 days of age, and continues to be expressed in the adult. Slot blot analysis demonstrates that TRPM-2 is also induced in the epididymis between 7 and 14 days of age, although, as has been demonstrated by in situ hybridization, TRPM-2 mRNA is detectable in the epithelial cells in the head of the epididymis but is barely detectable in the midportion or tail regions. Northern analysis suggests that the size of the TRPM-2 transcript in the testis also changes during development. In the early stages of testicular development, the TRPM-2 transcript appears to be a broad band of approximately 1.5 kb, while the transcript in the adult appears to be approximately 1.8 kb in length. S1-nuclease protection assays suggest that this increase in size is not due to differential splicing of the first exon of TRPM-2/SGP-2 and most probably reflects a difference in the polyadenylation of the mRNA in the testis at different times during development.
Animals↗
Gene expression in renal growth and regrowth.
To elucidate the molecular events associated with postnatal and compensatory renal growth, the expression of growth/differentiation genes (c-fos, c-myc, c-H-ras, c-K-ras), a stress-related gene (HSP70) and a structural gene (collagen type IV, alpha 1 and 2) were examined. Northern analysis of messenger ribonucleic acid from the newborn mouse reveals high levels of expression of HSP70, c-H-ras, and c-K-ras during the first week of life. By day 40 HSP70-related and c-H-ras expression decreases somewhat, c-K-ras remains unchanged and collagen type IV, which encodes for the renal glomerular basement membrane, expression decreases significantly. During compensatory renal growth increased expression of HSP70, c-H-ras and c-K-ras occurs. The results seem to indicate that growth/differentiation genes may be necessary for continued cell growth (hypertrophy) in postnatal and compensatory renal growth, and that collagen type IV formation continues up through week 2 of postnatal growth consistent with the interval of glomerular basement membrane formation.
Animals↗
Cascade induction of c-fos, c-myc, and heat shock 70K transcripts during regression of the rat ventral prostate gland.
Castration of an adult male rat results in the rapid regression of the ventral prostate gland. Most of the acinar epithelial cells lining the ducts of the gland will die during the first 5 days after androgen withdrawal. The molecular events which accompany the death of these cells were studied by examining RNA isolated from ventral prostate glands of normal and from a sequential 24-h series of castrate rats. These RNAs were analyzed by Northern blot methods to quantify the expression of growth-related (c-fos, c-myc, and heat shock 70K) and cell maintenance (alpha-tubulin) genes during prostatic regression. Each of these genes showed a bimodal pattern of expression. Within the first few days after castration, transcript levels decline; whereas later, during the most active period of cell death, their expression was transiently induced. Levels of mature c-myc, and alpha-tubulin transcripts increased approximately 6- to 8-fold on the third day after castration, while transcripts encoding hsp 70-related genes increased greater than 6-fold on the fourth day after castration. Further analysis of RNA extracted from regressing ventral prostate glands at sequential 12-h intervals after castration showed that the c-fos gene was induced at 36 h, earlier than c-myc, alpha-tubulin, and hsp 70.(ABSTRACT TRUNCATED AT 250 WORDS)
Androgens↗
Programmed cell death: early changes in metamorphosing cells.
Programmed cell death in intersegmental muscles and labial glands of Manduca sexta is a type II form of active cell death, in which lysosomal destruction of cytoplasm is a prominent and early feature, and the collapse of the nucleus is late and relatively modest until the bulk of the cytoplasm has been eroded. The prominent features of this collapse include an early expansion of the lysosomal compartment and a decrease in overall protein synthesis, while a small number of mRNAs persist or are upregulated. Energy resources appear to be adequate during the early stages of degeneration, and changes in levels of second messages likewise do not seem to be sufficiently dramatic to explain the failure of the tissue. There is some suggestion that DNA may not be completely intact long before the labial gland finally collapses, suggesting that it may fail under heavy load. Otherwise, we do not yet have an explanation for the precipitous drop in overall protein synthesis.
Animals↗
Metabolic events during programmed cell death in insect labial glands.
The labial gland of Manduca sexta is a valuable system to study the mechanisms of programmed cell death since the death of the gland is nearly synchronous and, except for the anterior duct, involves all of the tissue. The gland degenerates in 5 days during pupation. Our previous work documents a drop in total protein synthesis as the gland degenerates. To evaluate potential causes of this altered protein synthesis, we monitored several parameters of metabolism in dying cells: levels of adenosine triphosphate to estimate the energy resources of the gland; reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide to assess mitochondrial respiration; levels of acid phosphatase to assay lysosomal enzyme activity; and concentrations of cyclic nucleotides and inositol triphosphate to monitor signaling. While protein synthesis fell precipitously on day 0, total adenosine triphosphate and mitochondrial respiration were unchanged until the cells underwent massive collapse on day 3. Lysosomal acid phosphatase increased during early metamorphosis, and ultimately the bulk of the cytoplasm was destroyed in autophagic vacuoles. Changes in the concentrations of second messengers were modest and late. The relationships between the metabolism and the collapse of the labial gland are under investigation.
Acid Phosphatase↗