Liver Albumin mRNA Research Articles

Abolishment of proximal tubule albumin endocytosis does not affect plasma albumin during nephrotic syndrome in mice

The megalin/cubilin receptor complex is required for proximal tubular endocytosis and degradation of filtered albumin. An additional high-capacity retrieval pathway of intact albumin for the recovery of large amounts of filtered albumin has been proposed, possibly involving cooperation between megalin/cubilin and the neonatal Fc receptor. To clarify the potential role of such a pathway, we examined the effects of megalin/cubilin gene inactivation on tubular albumin uptake and plasma albumin levels in nephrotic, podocin knockout mice. Immunofluorescence microscopy of megalin/cubilin/podocin knockout mouse kidneys demonstrated abolishment of proximal tubule albumin uptake, in contrast to the excessive albumin accumulation observed in podocin knockout mice compared to controls. Correspondingly, urinary albumin excretion was increased 1.4 fold in megalin/cubilin/podocin compared to podocin knockout mice (albumin/creatinine: 226 vs. 157 mg/mg). However, no difference in plasma albumin levels was observed between megalin/cubilin/podocin and podocin knockout mice, as both were reduced to approximately 40% of controls. There were no differences in liver albumin synthesis by mRNA levels and protein abundance. Thus, megalin/cubilin knockout efficiently blocks proximal tubular albumin uptake in nephrotic mice but plasma albumin levels did not differ as a result of megalin/cubilin-deficiency, suggesting no significance of the megalin/cubilin-pathway for albumin homeostasis by retrieval of intact albumin.

Participation of liver progenitor cells in liver regeneration: lack of evidence in the AAF/PH rat model

When hepatocyte proliferation is impaired, liver progenitor cells (LPC) are activated to participate in liver regeneration. We used the 2-acetaminofluorene/partial hepatectomy (AAF/PH) model to evaluate the contribution of LPC to liver cell replacement and function restoration. Fischer rats subjected to AAF/PH (or PH alone) were investigated 7, 10 and 14 days post-hepatectomy. Liver mass recovery (LMR) was estimated, and the liver mass to body weight ratio calculated. We used serum albumin and bilirubin levels, and liver albumin mRNA levels to assess the liver function. LPC expansion was analyzed by cytokeratin 19 (CK19), glutathione S-transferase protein (GSTp) immunohistochemistry and by CK19, CD133, transforming growth factor-β1 and hepatocyte growth factor mRNA expression in livers. Cell proliferation was evaluated by Ki67 and BrdU immunostaining. Compared with PH alone where LMR was ∼100% 14 days post-PH, LMR was defective in AAF/PH rats (64.1±15.5%, P=0.0004). LPC expansion was scarce in PH livers (0.5±0.4% of CK19+ area), but significant in AAF/PH livers (8.5±7.2% of CK19+), and inversely correlated to LMR (r2=0.63, P<0.0001). A quarter of AAF/PH animals presented liver failure (low serum albumin and high serum bilirubin) 14 days post-PH. Compared with animals with preserved function, this was associated with a lower LMR (50±6.8 vs 74.6±9.4%, P=0.0005), a decreased liver to body weight ratio (2±0.3 vs 3.5±0.6%, P=0.001), and a larger LPC expansion such as proliferating Ki67+ LPC covered 17.4±4.2% of the liver parenchyma vs 3.1±1.5%, (P<0.0001). Amongst those, rare LPC with an intermediate hepatocyte-like phenotype were seen. Also, less than 2% of hepatocytes were engaged into the cell cycle (Ki67+), while more numerous (∼25% of hepatocytes) in the livers with preserved function. These observations suggest that, in this model, the efficient recovery of the liver function was ensured rather by the proliferation of mature hepatocytes than by the LPC expansion and differentiation into hepatocytes.

Effects of early postoperative enteral and parenteral nutrition support in cirrhotic rats after partial hepatectomy

Objective To evaluate the impact of early enteral and parenteral nutrition support in rats with liver cirrhosis after partial hepatectomy. Methods Twenty-four rats were randomly divided into three groups in this prospective randomized control study. From postoperative day ( POD) 1 to 5, all of these groups (group A, normal EN; group B, cirrhotic EN; group C, cirrhotic PN) after receiving partial hepatectomy were nutritionally supported with equal amount of calorie [690 kj/(kg·d)] and nitrogen [1.22 g/(kg·d)] content. Moreover, on the POD 6, parameters like weight, liver function, prealbumin and transferrin were tested, as well as level of liver albumin mRNA detected by reverse transcription-poly-merase chain reaction (RT-PCR) and morphological examinations, like HE stain and immunohistochemical stain assessed by index of Ki-67 protein. Results Compared with the preoperative baseline on the POD 6, group A gained weight, but not in the other groups [group A; (6.16 ±3.28) g; group B: (5.46 ±3.59) g; groupC; (6.13 ±3.21) g,P<0.05]. The levels of serum AST, ALT and ALP after partial hepatectomy were significantly higher in groups B and C than in group A (P <0.05). Serum level of ALP was higher in group C (271 ±92) IU/L than in group B [ (209 ± 81) IU/L.P <0.05 ]. Levels of serum prealbumin and transferring were significantly declined after partial hepatectomy in groups B and C as compared with group A (P<0.05). The levels of prealbumin were significantly higher in group B (161 ±37) mg/L than in group C [ (133 ±29) mg/L.P <0.05 ]. The levels of albumin mRNA and index of Ki-67 in the murine liver were compared among these groups. Albumin mRNA was suppressed in groups B and C as compared with group A, as well as that in group C as compared with group B (A: 1.46 ±0.12; B: 1.07 ±0.14; C; 0.84 ± 0.18, P < 0.05). Index of Ki-67 was higher in group A than in groups B and C, as well as that in group B than in group C (A: 8.90 ±0.45; B: 5.60 ±0.34; C: 3.80 ±0.21 ,P <0.05). Conclusion Active nutrition support, through EN nor PN cannot efficiently promote liver regeneration and relieve liver malfunction in cirrhotic rats rather than in normal ones after partial hepatectomy, represented by delayed restoration of various parameters reflecting liver function. Nevertheless, compared with PN, standard EN revealed some advantages though. Besides reducing risk of PN-related cholestasis, EN can better the restoration by promoting protein synthesis and tissue regeneration in the postoperative liver. Key words: Liver cirrhosis; Hepatectomy; Enteral nutrition; Hepatic regeneration

Synthesis rate of plasma albumin is a good indicator of liver albumin synthesis in sepsis.

Plasma albumin is well known to decrease in response to inflammation. The rate of albumin synthesis from both liver and plasma was measured in vivo by use of a large dose of L-[(2)H(3)-(14)C]valine in rats injected intravenously with live Escherichia coli and in pair-fed control rats during the acute-phase period (2 days postinfection). The plasma albumin concentration was reduced by 50% in infected rats compared with pair-fed animals. Infection induced a fall in both liver albumin mRNA levels and albumin synthesis relative to total liver protein synthesis. However, absolute liver albumin synthesis rate (ASR) was not affected by infection. In plasma, albumin fractional synthesis rate was increased by 50% in infected animals compared with pair-fed animals. The albumin ASR estimated in the plasma was similar in the two groups. These results suggest that hypoalbuminemia is not due to reduced albumin synthesis during sepsis. Moreover, liver and plasma albumin ASR were similar. Therefore, albumin synthesis measured in the plasma is a good indicator of liver albumin synthesis.

The molecular basis of Chinese herbs (Astragali and Angelica) on increasing serum albumin synthesis in nephrotic rats

SUMMARY: Our previous studies have shown that two Chinese herbs, Astragali and Angelica, combined with an adequate protein intake can ameliorate the reduction in serum albumin, and improve the ratio of protein synthesis to catabolic rate without significant influence on urinary protein in both nephrotic patients and rats. This study was designed to investigate mechanisms by studying the effect of Astragali and Angelica on albumin synthesis in experimental nephrotic syndrome (NS), measuring the effect on hepatic albumin mRNA expression and gene transcription. Male Sprague‐Dawley (SD) rats were divided into eight groups: (1) normal control, C; (2) nephrotic control, NC; (3) treated with Astragali and Angelica, Ast &amp; Ang; (4) treated with Astragali alone, Ast; (5) treated with Angelica alone, Ang; (6) treated with Astragali polysaccharide I, Ast I (MW. 36 000); (7) treated with Astragali polysaccharide II, Ast II (MW. 12 500); (8) treated with Astragali glucoside, Ast G. Nephrotic syndrome in rats was induced by accelerated nephrotoxic nephritis. the herb extracts were force‐fed daily for 14 days, the animals were then sacrificed, and the serum and liver collected. the level of serum albumin, albumin mRNA and its gene transcription activity were measured using biochemistry, Northern blot analysis, run‐on assay and quantity in laser density screening. In the NC group, the level of serum albumin was significantly lower, the level of albumin mRNA in the liver was significantly higher and the activity of albumin gene transcription was increased compared with normal controls. In the Ast &amp; Ang group, the urinary protein was not different to that in the NC group, while the level of serum albumin was significantly higher and both the level of albumin mRNA and the activity of albumin gene transcription were significantly higher. In each component therapy group, the effect of increasing the level of serum albumin, the levels of albumin mRNA and activities of albumin gene transcription were no different compared with the NC group. Our results suggest that Astragali and Angelica increase albumin synthesis in the nephrotic rat by increasing liver albumin mRNA expression, at least in part by improving the activity of gene transcription. the extract of Astragali and Angelica was better effect than any components tested.

In situ hybridization demonstration of albumin mRNA in B6C3F1 murine liver and hepatocellular neoplasms.

In situ hybridization was used to detect albumin mRNA in normal liver and hepatocellular neoplasms in 20 male B6C3F1 mice between 17 and 24 months of age. Positive signals for albumin were observed consistently in the cytoplasm of hepatocytes in normal liver, particularly in periportal areas. No signals were observed in other cells, such as Kupffer's cells, mesenchymal cells, or bile duct epithelium. Of hepatocellular adenomas, 11/11 (100%) stained positively for albumin mRNA, whereas 14/15 (93%) of primary hepatocellular carcinomas showed positive expression. Albumin mRNA was also detected in extrahepatic metastases of hepatocellular carcinoma, including 9/15 (60%) of pulmonary neoplasms and 5/12 (42%) of metastases at other sites. The pulmonary metastases of hepatocellular carcinoma frequently exhibited a glandular, papillary, or sarcomatous histologic appearance. The presence of albumin in these tumors, lacking characteristic hepatocellular phenotype, is a potential determinant of hepatic lineage. We conclude that in situ hybridization for albumin mRNA in mice is a useful tool in the differential diagnosis of hepatocellular carcinoma, particularly in the case of pulmonary metastasis. This technique may also enable recognition of hepatocyte differentiation in glandular structures with phenotypic features of biliary cells, as seen in mixed hepatocellular-cholangial neoplasms.

Insulin and dietary fructose induce stearoyl-CoA desaturase 1 gene expression of diabetic mice.

The transcription and mRNA levels of murine liver stearoyl-CoA desaturase 1 (SCD1) are induced 11- and 45-fold, respectively, by feeding fasted normal mice with a fat-free, high carbohydrate diet (Ntambi, J. M. (1992) J. Biol. Chem. 267, 10925-10930). In this study, we used streptozotocin-induced diabetic mice to study the regulatory role of carbohydrate and insulin on expression of the SCD1 gene in liver. Fructose administration to fasted diabetic mice induced a 2-fold increase in SCD1 mRNA within 6 h and a 23-fold increase within 24 h. Similarly, insulin administration to diabetic mice induced SCD1 mRNA from 4-fold within 4 h to 22-fold within 24 h. Insulin plus fructose, however, achieved full induction, with a 45-fold increase of SCD1 mRNA and a 10-fold increase in SCD1 transcription within 24 h. Additionally, the effect of insulin on SCD1 mRNA was inhibited 75% with dibutyryl-cAMP and theophylline administration and 70% by cycloheximide administration. Synthesis of liver albumin mRNA showed little change upon dietary manipulation or insulin treatment. Our data demonstrate that insulin and dietary fructose or a metabolite of fructose positively regulate the expression of the SCD1 gene in mouse liver.

In situ hybridization of albumin mRNA in normal liver and liver tumors: identification of hepatocellular origin.

In situ hybridization was performed to detect albumin mRNA in normal liver, liver cirrhosis, primary liver tumors and secondary liver neoplasms. In areas of normal liver, and liver cirrhosis, signals for albumin mRNA were present in hepatocytes, whereas no signals were seen in other cells such as endothelial and Küpffer cells, bile duct epithelium and smooth muscle cells. In 53 of 56 hepatocellular carcinomas signals were present in tumor cells but in eight cholangiocarcinomas and 14 metastatic adenocarcinomas from large bowel or pancreas, carcinoma cells were negative for albumin mRNA. In three metastatic tumors (from two neuroendocrine carcinomas and one gastric leiomyosarcoma), tumor cells contained no signals, while the surrounding hepatocytes showed diffuse grains. In 15 of the 84 specimens examined in situ hybridization was applied to routine formalin-fixed and paraffin-embedded blocks and strong signals were obtained for albumin mRNA. We conclude that in situ hybridization of human albumin is a valid tool in the differential diagnosis of hepatocellular carcinoma from cholangiocarcinomas and tumors metastatic to the liver.

Dietary regulation of stearoyl-CoA desaturase 1 gene expression in mouse liver.

Expression of the murine liver stearoyl-CoA desaturase gene (SCD1) is induced upon feeding fasted mice a fat-free, high carbohydrate diet (Ntambi, J. M., Buhrow, S. A., Kaestner, K. H., Christy, R. J., Sibley, E., Kelly, T. J., and Lane, M.D. (1988) J. Biol. Chem. 263, 17291-17300). In the present study, SCD1-specific RNA riboprobes and cDNA probes were used to study the mechanism for the induction of SCD1 mRNA. Based on the time course of induction, the SCD1 mRNA increased from 2-fold within 6 h to 45-fold within 36 h. Nuclear run-on transcription studies showed that the accumulation of SCD1 mRNA after refeeding starved mice a fat-free, high carbohydrate diet was a consequence of the transcriptional activation of the SCD1 gene. The SCD1 mRNA level decreased rapidly (t1/2 = approximately 4 h) within 24 h when mice fed the fat-free, high carbohydrate diet were switched to a regular chow diet. Furthermore, when the fat-free diet was supplemented with triacylglycerides containing polyunsaturated fatty acids, the transcription of the SCD1 gene and the induction of the SCD1 mRNA were significantly blunted. Triacylglycerides containing saturated and monounsaturated fatty acids were without dramatic effects. In contrast, synthesis of liver albumin mRNA was little affected by any of these dietary variations, indicating that the observed changes in the transcription of the SCD1 gene and mRNA levels were specific. These data demonstrate that both dietary carbohydrates and polyunsaturated fatty acids or their metabolites directly or indirectly regulate the expression of the SCD1 gene in mouse liver.

Modifications of hepatic alpha‐1‐acid glycoprotein and albumin gene expression in rats treated with phenobarbital

The serum level of alpha 1-acid glycoprotein (alpha 1-AGP) is significantly increased in various animal species by treatment with cytokines, glucocorticoids and phenobarbital. The mechanisms responsible for the cytokine-induced and glucocorticoid-induced increases are now well documented, but not so in the case of phenobarbital. The main purpose of this study was to assess whether phenobarbital acts on alpha 1-AGP synthesis in the liver at the transcriptional or translational level. Male Dark Agouti rats received 70 mg phenobarbital/kg daily for 7 days. The analysis of total hepatic RNA showed that a single injection of phenobarbital induced an 11-fold increase in phenobarbital-dependent cytochrome P450IIB mRNA, whereas seven injections of phenobarbital were required to induce a maximum 5.5-fold increase in alpha 1-AGP mRNA. Concurrently, the transcription rate of the alpha 1-AGP gene rose 3.5-fold. Hepatocytes isolated after the seventh injection of phenobarbital showed a threefold increased capacity to secrete alpha 1-AGP, corresponding to a 3.2-fold increased alpha 1-AGP mRNA content in the liver. In conditions in which its effect on the induction of alpha 1-AGP synthesis was maximum, phenobarbital caused a 30% reduction in liver albumin mRNA and in albumin secretion by isolated hepatocytes, resulting from a 60-70% reduction in the rate of transcription of the albumin gene measured in isolated nuclei. We conclude that the effect of phenobarbital on alpha 1-AGP and albumin gene expression occurs at the transcriptional rather than the translational level.

In situ hybridisation of albumin mRNA in normal liver and hepatocellular carcinoma with a digoxigenin labelled oligonucleotide probe.

To study the localisation and distribution of albumin mRNA in normal liver and hepatocellular carcinoma by in situ hybridisation with an oligonucleotide probe. A 51 base oligonucleotide was synthesised from a sequence at the 5' end of the human albumin gene and the probe was labelled at its 3' end with digoxigenin 11-dUTP. Formalin fixed, wax embedded sections of liver biopsy specimens were used to study the localisation and distribution of albumin mRNA. After in situ hybridisation the bound probe was visualised using a digoxigenin antibody conjugated with alkaline phosphatase. In normal liver albumin mRNA was detected in hepatocytes and no positive signal was observed in biliary epithelium, vascular endothelium, or Kupffer cells. In 75% (9/12) of the hepatocellular carcinomas studied a positive hybridisation signal was observed in tumour cells. Albumin mRNA can be detected in sections of formalin fixed, wax embedded liver, a digoxigenin labelled probe is ideally suited for in situ hybridisation of liver because there is no background from the detection system. The identification of albumin mRNA may be a useful marker of hepatocellular carcinoma, and the demonstration of albumin mRNA by in situ hybridisation overcomes the potential background problem associated with albumin immunohistochemistry.

Pretranslational regulation of albumin synthesis in tumor-bearing mice: The role of anorexia and undernutrition

Hepatic albumin synthesis, serum albumin turnover, and hepatic albumin messenger RNA (mRNA) content were evaluated in mice bearing a transplantable low differentiated tumor (MCG 101). Results obtained on tumor-bearing mice were compared with results obtained from non-tumor-bearing animals that were either freely fed, food restricted so that their body composition was similar to tumor-bearing animals (pair-weighed), fed a protein-free diet for 5 days, or fasted for 48 hours. Tumor-bearing animals became hypoalbuminemic (33 ± 5 vs. 44 ± 3 g/L in freely fed mice), which could be explained by both depressed albumin synthesis (1.95% ± 0.20% vs. 2.67% ± 0.27%/h in freely fed mice) and increased albumin degradation. Pair-weighed and protein-calorie malnourished controls had reductions in albumin synthesis (1.81% ± 0.18% and 1.67% ± 0.17%/h, respectively) similar to tumor-bearing animals, and the starved controls had the lowest synthetic rates (1.07% ± 0.10%/h). Albumin degradation was increased only in tumor-bearing animals. Hepatic albumin mRNA in undernourished animals was less (tumor bearing, 32% ± 5%; pair weighed, 47% ± 4%; 48 hours fasted, 18% ± 2%; and protein-calorie malnourished, 26% ± 3%) than 50% of the mRNA content in the livers of freely fed control mice. Messenger RNA-directed synthesis of albumin in vitro was also depressed to a variable degree in tumor-bearing and malnourished non-tumor-bearing controls. The hypoalbuminemia in tumor-bearing animals could not be prevented by daily injections of a prostaglandin synthesis inhibitor (indomethacin, 1 μg/g body wt), but the hepatic acute phase protein serum amyloid P decreased from 157 ± 12 to 103 ± 9 μg/mL in indomethacin-treated tumor-bearing mice (P < 0.01). It is concluded that increased albumin degradation seen in tumor-bearing animals cannot be explained by associated malnutrition, whereas tumor-associated malnutrition can explain to a large extent the depressed albumin synthesis. Decreased albumin synthesis in tumor-bearing animals correlated in part with a decreased quantity of liver albumin mRNA. The results of the current study are consistent with either a reduced transcription of the albumin gene or a change in albumin mRNA processing and stability communicated by anorexia and malnutrition.

A high-glucose, fat-free diet rapidly causes a specific decrease in albumin mRNA even if the diet contains adequate protein

We have shown previously that a high-glucose, fat-free diet decreases the level of liver albumin mRNA in rat even though the diet contains excess protein (30%, w/w). This diet is unique in that it also decreases the higher-order coiling of liver nuclear chromatin. Altered chromatin structure is considered a prerequisite for gene expression. In this study we measured the rate at which liver albumin mRNA is decreased by a high-glucose, fat-free diet. We compared the rate of reduction of albumin mRNA with the rate of dietary induction of two other prominent liver mRNAs. Using RNA-cDNA hybridization, we measured the relative amounts of mRNA for liver albumin, fatty acid synthetase (FAS), and 6-phosphogluconate dehydrogenase (6-PGD) after refeeding a high-glucose, fat-free diet for two hours to 5 days following a 48-hour fast. The results show that a high-glucose, fat-free diet which contains adequate protein causes a maximal decrease in albumin mRNA after 8 hours, coinciding with increases in the levels of FAS and 6-PGD mRNAs. The latter two mRNAs are required for the biosynthesis of fatty acids from the excess carbohydrate. The results indicate that the hepatocyte can coordinately regulate the levels of specific mRNAs, by down-regulating the level of albumin mRNA. The relatively long half-life of the albumin protein may be a factor in the regulation of its mRNA.

Molecular aspects of age-related changes in albumin synthesis in female WAG/Rij rats

The rate of albumin gene transcription and the level of albumin RNA sequences in liver nuclei of rats of various ages were determined to investigate the regulation of age-related changes in levels of cytoplasmic albumin mRNA in rat liver. No change was observed with age in both the rate of albumin gene transcription and the amount of nuclear albumin RNA sequences, which suggests that the increase in cytoplasmic albumin mRNA content in the rat liver with age is caused by a decreased turnover of this messenger. The length of poly(A)-tails of rat liver cytoplasmic RNA with age was also examined. Although no change with age was observed in the poly(A)-tail length sedimenting at the peak fraction, a shift towards shorter poly(A)-tails was found with age in the overall poly(A)-tail length distribution.

Low Protein—High Energy Diet Induces Repressed Transcription of Albumin mRNA in Rat Liver

The effects of diet at the molecular level were investigated by feeding rats control or protein-deficient diets. Each of the control and the protein-deficient groups was further divided into three subgroups according to the level of energy intake. Liver DNA, RNA and total cellular protein concentrations and serum albumin and albumin mRNA concentrations were determined. Protein deficiency caused a marked inhibition of liver growth but the size of most cells remained normal. The low protein diet caused concurrent decreases in the levels of serum albumin and serum albumin mRNA. However, this effect was observed only with the combination of a low protein diet and normal energy intake. The low protein-low energy diet failed to induce the low serum albumin level. Our findings suggest that the altered balance between at least two factors of protein and energy intake serves as a trigger for several metabolic changes which ultimately regulate specific genes.

Study of the molecular mechanism of decreased liver synthesis of albumin in inflammation.

Hypoalbuminemia in inflammatory disorders is not an infrequent finding. However, little is known about albumin synthesis in these patients. In the present study we have measured the albumin synthesis in four patients with inflammatory diseases using the [14C]carbonate technique. Because inflammation causes a decreased albumin synthesis and this decreased synthesis could not be related to a reduced amino acid supply, we have also examined the possible molecular mechanisms of reduced albumin synthesis during inflammation using in vivo and in vitro experiments in rats. In rats with turpentine-induced inflammation, serum albumin concentration and liver albumin mRNa level were markedly decreased. These changes could not be reproduced by administration of fibrinogen-, or fibrin-degradation products, or several hormones, such as corticosteroids, growth hormone, and adrenaline. However, monocytic products, especially interleukin 1, postulated to be important mediators of the inflammatory response, reduced albumin synthesis and liver albumin messenger RNA content but not total protein synthesis in rats in vivo and in primary cultures of rat hepatocytes. These findings suggest that monocytic products play an important role in reduced albumin synthesis during inflammation.

Changes in plasma albumin concentration, synthesis rate, and mRNA level during acute inflammation.

Induction of acute inflammation in rats by the subcutaneous injection of turpentine resulted in a marked fall in the concentration of albumin in plasma. This fall, which reached a minimum of 45% of the control level at 36 h after injection of the inflammatory agent, occurred in the presence of no significant change in the concentration of total protein in plasma. It was accompanied by a corresponding decline in the relative abundance of albumin mRNA in liver, which reached a minimum of 25% of the control level at 36 h after initiation of the inflammatory response. Perfused livers from 36-h postinjection rats exhibited albumin secretion rates that were reduced to 38% of control values. In contrast, release of total secretory proteins, secretion of nonalbumin plasma proteins, and synthesis of nonexported proteins by perfused livers were elevated to 166, 266, and 117% of the control values, respectively, as a result of the inflammatory response. These results demonstrate that acute inflammation causes a relative reduction in hepatic albumin mRNA, which leads to a corresponding decrease in albumin synthesis and secretion by liver and a fall in the concentration of albumin in plasma. The concentration of total protein in plasma is maintained during acute inflammation in part by increased synthesis and secretion by liver of nonalbumin plasma proteins, e.g. the acute-phase reactants.

Hepatic Level of Rat Albumin Messenger RNA is Influenced by Factors other than Dietary Protein

Dot hybridization of messenger RNA (mRNA) and complementary DNA (cDNA) has been used to measure the relative levels of albumin and α-fetoprotein mRNA in liver of rats fed for 5 d a fat-free (carbohydrate-rich) diet, a high fat diet or a basal diet, all three of which were isonitrogenous. The level of albumin mRNA in rats fed the fat-free (carbohydrate-rich) diet was 30 to 45% of the level in animals fed the basal 4% fat diet. The concentration of another mRNA, that for α-fetoprotein, remained unchanged. It has been established by others that albumin mRNA levels and albumin synthesis are diminished in response to low levels of dietary protein. We show that albumin mRNA levels are lower than those observed in animals fed the basal 4% fat diet, even when dietary protein is adequate (30% wt/wt), if the nonprotein calories are derived solely from carbohydrate.

Construction and cloning of rat albumin structural gene sequences.

A recombinant plasmid containing a DNA segment complementary to rat liver albumin mRNA has been constructed, cloned, and used to examine the organization of albumin gene. The 18S fraction of total liver poly(A)-containing RNA was copied into a double-stranded cDNA by avian myeloblastosis virus reverse transcriptase and Escherichia coli DNA polymerase I. The cDNA was inserted into the HindIII site of the plasmid pBR322 via the addition of specific oligonucleotide linkers. Recombinant plasmids were screened by hybrid arrest of mRNA translation and hybridization with specific cDNAs. Thereby, a plasmid was identified that contained a 1200-nucleotide insert corresponding to a segment adjacent to the 5'-terminal region of albumin mRNA. The inserted sequence was used as a hybridization probe to detect five EcoRI fragments of genomic DNA which encode albumin mRNA. These were compared to eight EcoRI fragments identified within the rat genome by albumin cDNA. We conclude that the albumin gene (or genes) is interrupted at more than one site in the coding DNA by intervening sequences. Furthermore, we were able to distinguish those fragments that encode the 5' and 3' ends of the mRNA.

Mouse albumin mRNA in liver and a hepatoma cell line. Preparation of complementary DNA from purified mRNA and quantitation by nucleic acid hybridization.

Mouse albumin mRNA in liver and a hepatoma cell line. Preparation of complementary DNA from purified mRNA and quantitation by nucleic acid hybridization.