Biomedical subjects

B T Kren

Publications and source records attributed to B T Kren.

42 records · Page 3Linked to original sources

PubMed1994

Posttranscriptional regulation of cyclin B messenger RNA expression in the regenerating rat liver.

The growing family of cyclin genes and their products have been identified as important regulatory participants in the eukaryotic cell cycle. Cyclin proteins are currently postulated to act at the G1 restriction point, entry and exit of S phase, and the G2-M transition. We have cloned a rat cyclin B complementary DNA (cDNA) and have investigated cyclin B mRNA expression and regulation in the regenerating rat liver following 70% partial hepatectomy (PH). Sequence analysis of the rat cyclin cDNA revealed greater than 82% identity to type B1 human and murine cyclin genes. The rat cyclin cDNA was used to probe Northern blots of polyadenylated enriched RNA from regenerating rat liver from 0 through 96 h post-PH. Two species of rat cyclin B transcript were detected which mapped at 1.6 and 2.4 kilobases in length. Steady-state transcript levels began to appear around 24 h post-PH, which coincides with peak DNA synthesis. However, expression of the cyclin B transcripts peaked at 48 h and was 20-fold greater than at 24 h post-PH. Smaller peaks of expression occurred at 30 and 72 h. Run-off transcription assays using nuclei isolated at various times post-PH indicated no change in transcriptional rate during the period of regeneration. In vivo mRNA half-life determinations were performed at 24, 40, and 48 h post-PH. The half-lives of both transcript species were almost identical and were determined to be greater than 12 h at 24 h post-PH, and 2.4 h at 40 and 48 h post-PH. Protein inhibition with cycloheximide increased the signal intensity of both transcripts between 48 and 54 h post-PH but had no detectable effect on 0 h transcript expression. Steady-state levels of thymidine kinase mRNA showed a similar pattern of expression by Northern analysis through 96 h post-PH as cyclin B. The present study indicates that the appearance of cyclin B mRNA in the regenerating rat liver is coincident with peak DNA synthesis, although its own peak expression is significantly delayed. Steady-state transcript levels appear to be regulated primarily by posttranscriptional events of which changes in mRNA stability may be an important determinant. We propose that the involvement of cyclin B in the cell cycle machinery is controlled at several different levels of gene expression.

Amino Acid Sequence↗

PubMed1994

Transcriptional rate and steady-state changes of retinoblastoma mRNA in regenerating rat liver.

This study characterizes the mRNA expression of the retinoblastoma tumor suppressor gene in regenerating rat liver during 96 hr after 70% partial hepatectomy. A 960-bp BglII-OxaNI fragment of murine retinoblastoma cDNA was used to probe Northern blots of poly(A)(+)-enriched RNA isolated from regenerating liver. Two species of retinoblastoma mRNA, 2.8 kb and 4.7 kb long, were identified in control liver and exhibited an intensity ratio of 5:1, respectively. Expression of the 2.8-kb mRNA was reduced by 50% 1 hr after partial hepatectomy and was less than 10% of control values by 3 hr. The transcript began to reappear at 12 hr and returned to near-baseline levels by 24 hr. In contrast to the rapid disappearance of the 2.8-kb transcript, expression of the 4.7-kb mRNA increased 15-fold by 6 hr and returned to control levels by 18 hr after partial hepatectomy. Pretreatment of the animals with cycloheximide before partial hepatectomy completely stabilized steady-state levels of both mRNA transcripts through 6 hr. Nuclear run-on assays revealed a sixfold increase in transcription by 30 min and a return to near-baseline levels by 6 hr. The in vivo half-lives of the 2.8- and 4.7-kb transcripts in control livers were 39.5 and 41.2 min, respectively. The half-life of the 4.7-kb transcript 6 hr after partial hepatectomy was 39.1 min. Intravenous administration of transforming growth factor-beta 1, a known inhibitor of hepatocyte replication, just before partial hepatectomy caused no significant change in the modulation of the transcripts through 24 hr.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Differential regulation of multiple gap junction transcripts and proteins during rat liver regeneration.

The mRNA and protein expression of alpha 1 (connexin 43), beta 1 (connexin 32), and beta 2 (connexin 26) gap junction genes were examined in the regenerating rat liver after 70% partial hepatectomy (PH). Expression of beta 1 and beta 2 steady-state mRNA levels changed minimally until 12 h after PH when both transcripts decreased to approximately 15% of baseline values. A similar decrease in assembled connexin levels was detected by immunoblot and indirect immunofluorescence at 18 h after PH. Both transcripts simultaneously increased between 24 and 42 h and again rapidly decreased by 48 h post-PH. beta 1 and beta 2 assembled gap junction protein expression increased at 48 h post-PH and rapidly decreased by 56 h. By 72 to 84 h post-PH, beta 1 and beta 2 mRNA and assembled protein expression returned to near baseline levels and were maintained. Interestingly, inhibition of protein synthesis with cycloheximide completely inhibited disappearance of the beta 2 transcript, in contrast to beta 1 mRNA which was unaffected. Nuclear run-on assays showed no change in transcriptional rates for either gene during the regenerative period. However, both beta 1 and beta 2 transcripts exhibited significantly decreased mRNA half-lives at 12 h post-PH (3.8 and 3.7 h, respectively) relative to those at 0 h (10.9 and 6.1 h, respectively). Surprisingly, although the transcriptional rate for alpha 1 was similar to that observed for beta 2, no alpha 1 transcripts were detectable by northern or RNase protection analysis. The results suggest that in the regenerating rat liver, beta 1 and beta 2 gap junction genes are not regulated at the transcriptional level. Rather, the cyclical modulation of their steady-state transcripts is regulated primarily by posttranscriptional events of which mRNA stability is at least one critical factor in the control process.

Animals↗

PubMed1993

Cyclin and cyclin-dependent kinase 1 mRNA expression in models of regenerating liver and human liver diseases.

There is compelling evidence that the eukaryotic cell cycle is controlled by a family of proteins called cyclins, which complex with cyclin-dependent kinases (CDK) to modulate key events during cell division. We have examined the regulation of these genes in models of experimental liver regeneration and their expression in human liver diseases. Seventy percent partial hepatectomy (PH) was performed on rats and normal BALB/c and athymic nude mice to determine patterns of cyclin and CDK1 mRNA expression. It has been previously shown by [3H]thymidine incorporation that athymic nude mice manifest impaired regeneration after PH. Our results demonstrate a sequential pattern of cyclin and CDK1 transcript expression in each of the models. Cyclin D1 was the most abundant mRNA steady-state transcript in the regenerating livers. CDK1 and cyclins associated with later stages of the cell cycle showed delayed and diminished expression in nude mice compared with normals. Nuclear run-off assays performed at key time points post-PH revealed little change in transcription rates, suggesting that steady-state mRNA expression of the cyclin genes is regulated primarily by posttranscriptional events. Human liver tissue from various acute and chronic hepatic diseases showed increased expression of cyclins A and D1. We conclude that the regenerating liver post-PH offers an excellent in vivo model for studying cyclin and CDK gene expression. Impaired regeneration in the nude mouse is associated with altered cyclin and CDK1 mRNA transcript expression. Furthermore, cyclins may eventually provide clinically relevant molecular markers of regenerative activity in human liver diseases.

Adolescent↗

PubMed1991

Ethanol interferes with regeneration-associated changes in biotransforming enzymes: a potential mechanism underlying ethanol's carcinogenicity?

The effects of chronic ethanol consumption on enzyme systems involved in carcinogen activation and detoxification were studied in a rat model of liver regeneration. In control rats, steady-state messenger RNAs of cytochrome P450j decreased 12 to 24 hr after partial hepatectomy but were fully recovered by 48 to 72 hr. In contrast, messenger RNA levels of cytochrome P450b and P450d did not vary significantly during that period. Steady-state messenger RNA levels for the placental form of glutathione S-transferase decreased within 30 min after partial hepatectomy but fluctuated until levels returned to normal by 48 hr. Preliminary nuclear run-on analyses suggest that the regulation of cytochrome P450j and the placental form of glutathione S-transferase messenger RNA levels involves posttranscriptional control in these animals. In ethanol-fed rats, as in controls, expression of cytochrome P450j and the placental form of glutathione S-transferase decreased transiently after partial hepatectomy. However, compared with control values, messenger RNA levels for cytochrome P450j were greater in ethanol-fed rats at each time point. Similar results were noted for placental glutathione S-transferase levels from 12 to 48 hr after partial hepatectomy. Ethanol feeding had no apparent effect on steady-state messenger RNA levels of cytochrome P450d, P450b or the multidrug-resistant gene. In both ethanol and control rats, only prehepatectomy levels of cytochrome P450 transcripts correlated with levels of the respective P450 isoenzymes. These data indicate that liver regeneration selectively decreases the steady-state messenger RNA expression of certain isoenzymes of cytochrome P450 and glutathione S-transferase.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1980

Dominant lethality in Xenopus laevis induced with triethylenemelamine (TEM).

Adult male South African clawed toads, Xenopus laevis, were injected intraperitoneally with triethylenemelamine (TEM) dissolved in water over a dose range of 13-1,300 micrograms/kg or with water (controls). Mating of the treated males with untreated females was induced seven days later by injection of human chorionic gonadotropic hormone (HCG). Accumulated lethality one week after fertilization among progeny was dose related. Morphological abnormalities among embryos that survived to hatching were similarly dose related. Because anatomically anomalous embryos may have impaired behavior, swimming capacity was tested, and anomalies in swimming behavior were also found to be dose related. Finally, short-term cell cultures were made on minced embryos to obtain chromosome spreads. Structural aberrations, rings, dicentrics, etc of somatic chromosomes were also dose related.

Animals↗