Human Liver RNA Research Articles

Identification of a broadly fibrogenic macrophage subset induced by type 3 inflammation.

Macrophages are central orchestrators of the tissue response to injury, with distinct macrophage activation states playing key roles in fibrosis progression and resolution. Identifying key macrophage populations found in human fibrotic tissues could lead to new treatments for fibrosis. Here, we used human liver and lung single-cell RNA sequencing datasets to identify a subset of CD9+TREM2+ macrophages that express SPP1, GPNMB, FABP5, and CD63. In both human and murine hepatic and pulmonary fibrosis, these macrophages were enriched at the outside edges of scarring and adjacent to activated mesenchymal cells. Neutrophils expressing MMP9, which participates in the activation of TGF-β1, and the type 3 cytokines GM-CSF and IL-17A coclustered with these macrophages. In vitro, GM-CSF, IL-17A, and TGF-β1 drive the differentiation of human monocytes into macrophages expressing scar-associated markers. Such differentiated cells could degrade collagen IV but not collagen I and promote TGF-β1-induced collagen I deposition by activated mesenchymal cells. In murine models blocking GM-CSF, IL-17A or TGF-β1 reduced scar-associated macrophage expansion and hepatic or pulmonary fibrosis. Our work identifies a highly specific macrophage population to which we assign a profibrotic role across species and tissues. It further provides a strategy for unbiased discovery, triage, and preclinical validation of therapeutic targets based on this fibrogenic macrophage population.

Abstract 155: Mutations In ABCA8 Result In HDL Deficiency And Cholesterol Efflux Defects

The molecular basis of extreme HDL-C levels is incompletely understood. To identify mutations in novel putative HDL genes, we sequenced select genes in 80 unrelated individuals with extreme low HDLc (HDL %ile≤10th) and as controls, 120 individuals with high HDL (HDL %ile≥90th). We identified two variants in ABCA8 in subjects with low HDL-C levels. The E18-2T>C mutation is located in a highly conserved region and gives rise to a disruption of a splice site. The G>C variant occurs at a site conserved in vertebrates, at position 64425884 (build HG18), and results in Pro609Arg substitution. No ABCA8 mutations were identified in HHDL individuals. These represent the first mutations identified in ABCA8, which belongs to the family of ATP binding cassette transporters. Family expansion followed by genotyping identified additional mutation carriers and first degree relative controls in whom segregation analyses were performed. Compared with controls, mutation carriers showed a significant 32.6% decrease in plasma HDLc levels and decreased HDL percentiles (HDLc: controls = 1.23±0.26, mutation carriers = 0.83±0.37, mmol/L, p=0.0007; HDL%ile: controls = 41.8±21.2, mutation carriers = 17.3±19.2, p=0.004, n=23 controls and 12 carriers). No changes in LDLc, triglycerides or BMI were observed. Since mutations in ABCA8 reduce plasma HDLc levels, we assessed the role of ABCA8 in lipid efflux. Cholesterol efflux to lipid-free APOA-I was significantly reduced by 62.1% in fibroblasts from the proband with the E18-2 T>C mutation. In comparison, fibroblasts from heterozygous ABCA1 mutation carriers showed a 30-60% decrease in efflux to APOA-I depending on mutation severity. Wild type and P609R ABCA8 cDNA clones were generated from human liver RNA. A significant ~200% increase in cholesterol efflux to lipid free APOA-I was observed with wild type ABCA8, further suggesting that ABCA8 is a lipid transporter. A significant 75% decrease in the efflux of cholesterol to lipid-free APOA-I (p=0.02) was observed with the mutant P609R ABCA8 protein compared to wild-type. We show here that ABCA8 is a cholesterol transporter, and that mutations in ABCA8 are a novel cause for reduced plasma HDLc levels in humans.

Hepatitis C Virus Suppresses C9 Complement Synthesis and Impairs Membrane Attack Complex Function

Hepatitis C virus (HCV) proteins inhibit complement component expression, which may attenuate immunity against infection. In this study, we examined whether HCV regulates the membrane attack complex (MAC) via complement component C9. MAC is composed of C5b to C9 (C5b-9) and mediates cell lysis of invaded pathogens. Liver biopsy specimens from chronically HCV-infected patients exhibited a lower level of C9 mRNA expression than liver biopsy specimens from unrelated disease or healthy control human liver RNA. Hepatocytes infected with cell culture-grown HCV or expressing HCV core protein also displayed significant repression of C9 mRNA and protein levels. Promoter analysis suggested that the T cell factor-4 (TCF-4E) transcription factor is responsible for HCV core-mediated C9 promoter regulation. Sera from chronically HCV-infected patients displayed a lower level of C5b-9 and a reduced antimicrobial effect on model organisms compared to unrelated patient sera or sera from healthy volunteers. Together, these results for C9 regulation by HCV core protein coupled with functional impairment of the membrane attack complex underscore HCV-mediated attenuation of immune mechanisms.

Identification of microRNAs as a potential novel regulatory mechanism in HSD11B1 expression

11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1, gene HSD11B1) converts glucocorticoid receptor-inert cortisone to receptor-active cortisol. Multiple evidence supports a causal role for 11β-HSD1 in the current obesity epidemic. In obese, HSD11B1 expression is increased in adipose tissue but typically decreased in liver, and the underlying tissue-specific mechanisms are largely unknown. In this context, we investigated a potential role of microRNAs (miRNAs). We used several miRNA target prediction tools to identify possible candidates and a publicly available miRNA expression atlas to further select candidates expressed in hepatocytes. Using a dual luciferase reporter assay, we identified three potential miRNAs, hsa-miR-340, -561 and -579, as potential negative regulators of HSD11B1 expression. Disruption of the corresponding microRNA response elements abolished repression of luciferase activity for hsa-miR-561 and -579, but not for hsa-miR-340. Furthermore, levels of firefly luciferase mRNA were not changed by miR-561 and -579, indicating a mechanism based on translational repression rather than mRNA degradation. Finally, we were able to detect both, miR-561 and -579, in human total liver RNA by reverse-transcription-polymerase chain reaction (RT-PCR). According to the presented results, miR-561 and -579 are likely to be involved in the tissue-specific regulation of HSD11B1 expression. Moreover, literature findings and a pathway enrichment analysis support a potential role of these two miRNAs in glucocorticoid metabolism and signalling and associated diseases.

A homozygous nonsense mutation (c.214C→A) in the biliverdin reductase alpha gene (BLVRA) results in accumulation of biliverdin during episodes of cholestasis

BackgroundGreen jaundice is a rare finding usually associated with end-stage liver disease. ObjectiveThe authors investigated two unrelated Inuit women from different geographical areas in Greenland who had episodes of green...

Analysis of hepatic genes involved in the metabolism of fatty acids and iron in nonalcoholic fatty liver disease

Hepatic steatosis and iron cause oxidative stress, thereby progressing steatosis to steatohepatitis. We quantified the expression of genes involved in the metabolism of fatty acids and iron in patients with nonalcoholic fatty liver disease (NAFLD). The levels of transcripts for the following genes were quantified from biopsy specimens of 74 patients with NAFLD: thioredoxin (Trx), fatty acid transport protein 5 (FATP5), sterol regulatory element-binding protein 1c (SREBP1c), fatty acid synthase (FASN), acetyl-coenzyme A carboxylase (ACAC), peroxisome proliferative activated receptor alpha (PPARalpha), cytochrome P-450 2E1 (CYP2E1), acyl-coenzyme A dehydrogenase (ACADM), acyl-coenzyme A oxidase (ACOX), microsomal triglyceride transfer protein (MTP), transferrin receptor 1 (TfR1), transferrin receptor 2 (TfR2) and hepcidin. Twelve samples of human liver RNA were used as controls. Histological evaluation followed the methods of Brunt. The levels of all genes were significantly higher in the NAFLD patients than in controls. The Trx level increased as the stage progressed. The levels of FATP5, SREBP1c, ACAC, PPARalpha, CYP2E1, ACADM and MTP significantly decreased as the stage and grade progressed (P < 0.05). Hepatic iron score (HIS) increased as the stage progressed. The TfR1 level significantly increased as the stage progressed (P < 0.05), whereas TfR2 level significantly decreased (P < 0.05). The ratio of hepcidin mRNA/ferritin (P < 0.001) or hepcidin mRNA/HIS (P < 0.01) was significantly lower in NASH patients than simple steatosis patients. Steatosis-related metabolism is attenuated as NAFLD progresses, whereas iron-related metabolism is exacerbated. Appropriate therapies should be considered on the basis of metabolic changes.

Novel Polymorphic Human UDP-glucuronosyltransferase 2A3: Cloning, Functional Characterization of Enzyme Variants, Comparative Tissue Expression, and Gene Induction

UDP-glucuronosyltransferases (UGTs) are critical to the detoxification of numerous drugs, environmental pollutants, and endogenous molecules. However, as yet not all of the human UGTs have been cloned and characterized. cDNA clones from the UGT2A3 gene (located on chromosome 4q13) were isolated using pooled human liver RNA. Approximately 10% of clones contained a c.1489A>G nucleotide substitution, yielding proteins with a residue 497 alanine (UGT2A3.2) instead of a threonine (UGT2A3.1). The allele frequency of this polymorphism (rs13128286) was 0.13 in a European-American population as determined by direct DNA sequencing. Of 81 structurally diverse glucuronidation substrates tested, UGT2A3 expressed by a baculovirus system selectively glucuronidated bile acids, particularly hyodeoxycholic acid at the 6-hydroxy position. Apparent K(m) values of UGT2A3.1 and UGT2A3.2 for hyodeoxycholic acid 6-glucuronidation were 69 +/- 7 and 44 +/- 12 microM, respectively. Of 29 different extrahepatic tissues evaluated by real-time polymerase chain reaction, UGT2A3 mRNA was most highly expressed in small intestine (160% of liver), colon (78% of liver), and adipose tissue (91% of liver). An in silico scan of the proximal UGT2A3 promoter/5'-regulatory region identified transcription factor consensus elements consistent with tissue-selective expression in liver (HNF1) and intestine (CXD2), as well as induction by rifampicin (pregnane X receptor). In LS180 human intestinal cells, rifampicin increased UGT2A3 mRNA by more than 4.5-fold compared with vehicle, whereas levels were not significantly affected by the arylhydrocarbon receptor ligand beta-naphthoflavone. This is the first report establishing UGT2A3 as a functional enzyme, and it represents significant progress toward the goal of having a complete set of recombinant human UGTs for comparative functional analyses.

Expression of human CYP2E1 in insect cells using bac-to-bac expression system

To obtain recombinant human CYP2E1 and to determine its activity by using the specific probe substrate. CYP2E1 cDNA was obtained by RT-PCR using human liver RNA as template. The cloned CYP2E1 cDNA was ligated with pFastBac vector to generate recombinant pFastBac-CYP2E1, which was then transformed into E. coli DH 10 Bac. Recombinant Bacmid-CYP2E1 was generated by transposition. Then Spodoptera frugiperda (Sf9) insect cells was infected with Bacmid-CYP2E1 to generate recombinant baculoviruses carrying human CYP2E1 cDNA. Finally, Sf9 insect cells were triinfected with recombinant baculoviruses carrying human CYP2E1, CYPOR and CYPb5. The activity of the recombinant enzymes was determined using chlorzoxazone as the substrate. The Kmand Vmaxof recombinant CYP2E1 to chlorzoxazone was (72.4 +/-8.7) micromol. L(-1) and (2.41 +/-0.10) micromol.min(-1)?g(-1)protein, respectively. Active recombinant CYP2E1 has been obtained by bac-to-bac expression system and its activity is similar to previous reports.

TRAILs and tribulation

TRAILs and tribulation

Molecular Cloning and Characterization of Expressed Human Ecto-Nucleoside Triphosphate Diphosphohydrolase 8 (E-NTPDase 8) and Its Soluble Extracellular Domain

An ecto-nucleoside triphosphate diphosphohydrolase (ecto-NTPDase) has been cloned from human liver RNA by RT-PCR. The 1.5 kb cDNA codes for a protein of 495 amino acids. Sequence analysis indicated that it is most closely related to a chicken ecto-ATPDase previously cloned in our laboratory [Knowles et al. (2002) Eur. J. Biochem. 269, 2373-2382] and a mouse homologue that has been designated as E-NTPDase 8 [Bigonnesses et al. (2004) Biochemistry 43, 5511-5519]. The human E-NTPDase 8 has similar topology as the avian and mouse E-NTPDase 8 but has fewer potential N-glycosylation sites and only two amino acid residues in the cytoplasm at its C-terminus. Despite 52% identity in primary structures, enzymatic properties of human E-NTPDase 8 expressed in HEK293 cells differ from that of the chicken E-NTPDase 8. In contrast to the chicken E-NTPDase 8, the human E-NTPDase 8 hydrolyzes MgADP poorly and is inhibited by several detergents as well as benzyl alcohol; the latter attribute may be related to weaker interaction of the transmembranous domains of the human E-NTPDase 8. To demonstrate that inhibition by detergents is mediated by the transmembranous domains, a recombinant pSecTag2 plasmid containing the extracellular domain (ECD) of the human E-NTPDase 8 was constructed. The soluble human E-NTPDase 8 which was secreted into the culture media of transfected HEK293 cells was purified by ammonium sulfate fractionation and nickel affinity chromatography. Besides becoming resistant to detergent inhibition, the soluble human E-NTPDase 8 ECD displays greater activity with Ca nucleotide substrates, an increased affinity for ATP, different pH dependence, and a decreased sensitivity to azide inhibition when compared to the membrane-bound enzyme. These differences may result from the different conformations that the ECD assume without or with constraints exerted by the transmembranous domains. These results indicate that the transmembranous domains are important in regulating enzyme activity as well as in determining the structure of human E-NTPDase 8.

Inter-individual variation of several cytochrome P450 2D6 splice variants in human liver

To examine the possibility that inter-individual differences in splicing partially explain the observed differences in CYP2D6 activity, we amplified its full-length cDNA in 96 human liver RNA samples and discovered five splice variants: intron 5 retention, intron 6 retention, intron 5 and intron 6 double retention, exon 3 skipping, and partial intron 1 retention. All of the CYP2D6 splice variants we identified are probably nonfunctional transcripts. Substantial inter-individual variation in the proportions of the CYP2D6 transcript represented by splice variants, measured by real-time PCR, suggests that the presence of these splice variants contributes to the population variation in CYP2D6 activity. Relatively high levels of intron 6 retention were not correlated with the newly discovered single nucleotide polymorphism 2988G > A in intron 6 ( CYP2D6∗ 41) but did correlate with the more common CYP2D6∗ 34 allele. Our study prompts further investigations to explore the effect of these splice variants on drug metabolism.

IMPACT OF TRANSCRIPTION FACTOR PROFILE AND CHROMATIN CONFORMATION ON HUMAN HEPATOCYTE CYP3A GENE EXPRESSION

Recent data have made it increasingly clear that the gene expression profile of a cell system, and its alteration in response to external stimuli, is highly dependent on both the higher order chromatin structure of the genome and the interaction of gene products in interpreting stimuli. To further explore this phenomenon, we have examined the role of both of these factors in controlling xenobiotic-mediated gene expression changes in primary and transformed human hepatocytes (HuH7). Using quantitative polymerase chain reaction, expression levels of several transcription factors implicated in the liver-specific regulation of the CYP3A gene family were examined in human adult and fetal liver RNA samples. These expression profiles were then compared with those obtained from both primary and transformed human hepatocytes, showing that, in general, cultured cells exhibit a distinct profile compared with either the fetal or adult samples. Transcriptome profiles before and after exposure to the CYP3A transcriptional activators rifampicin, dexamethasone, pregnane-16alpha-carbonitrile, and phenobarbital were subsequently examined. Whereas exposure to these compounds elicited a dose-dependent increase in CYP3A transcription in primary hepatocytes, no alteration in expression levels was observed for the hepatoma cell line HuH7. Alteration in the expression levels of pregnane X receptor and chicken ovalbumin upstream promoter transcription factor I, and the disruption of higher order chromatin within HuH7 cells altered CYP3A expression and/or activation by xenobiotics toward that observed in primary hepatocytes. These data provide potential roles for these two processes in regulating CYP3A expression in vivo.

An RT-PCR-based Strategy to Estimate Full-Length CYP2D6 mRNA Copy Number

The goal of this study is to develop an analytical tool to assess cytochrome P450 2D6 (CYP2D6) levels in the form of full-length transcripts. CYP2D6 RNA in test samples was evaluated by co-amplification with an internal RNA control in a reverse-transcribed polymerase chain reaction (RT-PCR). The internal CYP2D6 RNA control was constructed by internally deleting 474 bp of CYP2D6 RNA, allowing simultaneous amplification of the test RNA together with the internal control RNA in a single RT-PCR reaction. With sequential dilution of test RNA, the CYP2D6 mRNA transcript levels in test samples were estimated. The full-length RT-PCR strategy allowed semiquantitative assessments of CYP2D6 RNA transcripts with a sensitivity limit of 500 copies for CYP2D6 RNA transcripts, 2500 copies/microg total human liver RNA, and 10% intraday coefficient of variation (CV). In a method validation study, the CYP2D6 activity appeared to relate more closely to full-length CYP2D6 mRNA concentration than a short-sequence of CYP2D6 RNA estimated with a real-time quantitative RT-PCR assay. We have developed an efficient semiquantitative assay and demonstrated its suitability for estimating full-length CYP2D6 mRNA transcripts in cells and tissues.

An intronic endogenous retrovirus-like sequence attenuates human haptoglobin-related gene expression in an orientation-dependent manner

The human haptoglobin-related gene ( HPR) gene codes for a haptoglobin-related protein (Hpr), a component of trypanosome lytic factor which circulates in plasma in small quantities. Except for the presence of a retrovirus-like element, RTVL-Ia, in intron 1, HPR is 92% identical in sequence to the closely linked human haptoglobin gene ( HP) gene coding for haptoglobin. We have explored experimentally in tissue culture and in vivo in mice and in humans the influence of the retroviral-like sequence type Ia (RTVL-Ia) element on HPR expression. Transient expression in HepG2 cells of plasmids carrying the HPR promoter joined by a shortened version of intron 1 to the chloramphenicol acetyltransferase (CAT) vector showed that fragments containing the 5′ long terminal repeat (LTR) had no significant effect. In contrast, a gag-pol related part and a pol-env-3′LTR related part of RTVL-Ia decreased expression to 20% and 40% of that in their absence but only when they were in naturally occurring orientation. The latter fragment that contains sequences reminiscent of elements essential for retrovirus viability, such as a splicing acceptor site, TATA box and polyA addition signal sequence, was further tested in site-specific transgenic mice. Similar to in vitro experiment, insertion of this fragment into an HPR transgene in mice reduced HPR expression to 50% compared to a transgene without the insert, but none of the viral sequence motifs appear to explain this effect. Instead, we found within the fragment two cryptic splicing donor sites whose products were present in transgenic mouse and in human liver RNA. Our data suggest that a combination of multiple small effects of RTVL-Ia including aberrant splicing accounts for the low (6%) expression of the present-day HPR relative to HP.

Cloning and characterization of human liver cytosolic β-glycosidase

Cytosolic β-glucosidase (EC 3.2.1.21) from mammalian liver is a member of the family 1 glycoside hydrolases and is known for its ability to hydrolyse a range of β-d-glycosides, including β-d-glucoside and β-d-galactoside. We therefore refer to this enzyme as cytosolic β-glycosidase. We cloned the cDNA encoding the human cytosolic β-glycosidase by performing PCR on cDNA prepared from total human liver RNA. Specific primers were based on human expressed sequence tags found in the expressed sequence tag database. The cloned cDNA contained 1407nt with an open reading frame encoding 469 amino acid residues. Amino acid sequence analysis indicates that human cytosolic β-glycosidase is most closely related to lactase phlorizin hydrolase and klotho protein. The enzyme was characterized by using cell lysates of COS-7 cells transfected with a eukaryotic expression vector containing the cDNA. The biochemical, kinetic and inhibition properties of the cloned enzyme were found to be identical with those reported for the enzyme purified from human liver.

Cloning and characterization of human liver cytosolic beta-glycosidase.

Cytosolic beta-glucosidase (EC 3.2.1.21) from mammalian liver is a member of the family 1 glycoside hydrolases and is known for its ability to hydrolyse a range of beta-D-glycosides, including beta-D-glucoside and beta-D-galactoside. We therefore refer to this enzyme as cytosolic beta-glycosidase. We cloned the cDNA encoding the human cytosolic beta-glycosidase by performing PCR on cDNA prepared from total human liver RNA. Specific primers were based on human expressed sequence tags found in the expressed sequence tag database. The cloned cDNA contained 1407 nt with an open reading frame encoding 469 amino acid residues. Amino acid sequence analysis indicates that human cytosolic beta-glycosidase is most closely related to lactase phlorizin hydrolase and klotho protein. The enzyme was characterized by using cell lysates of COS-7 cells transfected with a eukaryotic expression vector containing the cDNA. The biochemical, kinetic and inhibition properties of the cloned enzyme were found to be identical with those reported for the enzyme purified from human liver.

Characterization of the gene encoding human TAFI (thrombin-activable fibrinolysis inhibitor; plasma procarboxypeptidase B).

Thrombin-activable fibrinolysis inhibitor (TAFI) is a recently described human plasma zymogen that is related to pancreatic carboxypeptidase B. The active form of TAFI (TAFIa), which is formed by thrombin cleavage of the zymogen, likely inhibits fibrinolysis by removal from partially degraded fibrin of the carboxyl-terminal lysine residues which act to stimulate plasminogen activation. We have isolated and characterized genomic clones which encompass the entire human TAFI gene from lambda phage and bacterial artificial chromosome genomic libraries. The complete TAFI gene contains 11 exons and spans approximately 48 kb of genomic DNA. The positions of intron/exon boundaries are conserved between the TAFI gene and the rat pancreatic carboxypeptidase A1, A2, and B and the human mast cell carboxypeptidase A genes, indicating that these carboxypeptidases arose from a common ancestral gene. However, the intron lengths diverge significantly among all of these genes. The TAFI promoter lacks a consensus TATA sequence, and transcription is initiated from multiple sites. Transient transfection of reporter plasmids containing portions of the TAFI 5'-flanking region into mammalian cells allowed localization of the promoter and identified a approximately 70 bp region crucial for liver-specific transcription. Sequence analysis of cDNA clones obtained from human liver RNA indicated that the TAFI transcript is polyadenylated at three different sites. Our findings will facilitate the assessment of the regulation of TAFI expression by transcriptional and/or posttranscriptional mechanisms. Furthermore, knowledge of the genomic structure of the TAFI gene will aid in the identification of mutations that may be associated with the tendency to either bleed or thrombose.

Structure of the gene mutated in glycogen storage disease type Ib.

We report the structure of the human gene encoding the putative glucose 6-phosphate translocase that is mutated in glycogen storage disease type Ib. Northern blots showed that the encoded 2.4 kb mRNA is mainly expressed in liver and in kidney, but is also present, although in barely detectable amounts, in leucocytes. The gene contains nine exons, one of which (exon 7) is not present in human liver or leucocyte RNA. RT-PCR analysis of mouse RNA indicates that exon 7, which is 63 bp long compared with 66 bp in man, is not expressed in liver and kidney but well in heart and brain. 5'-RACE and RNase protection assays performed on RNAs from human liver, kidney and leucocytes indicated the presence of two main regions of transcription start at approximately -200 and -100 bp with respect to the initiator ATG.

Protective effects of extracted human-liver RNA, a known interferon inducer, against radiation-induced cytogenetic damage in male mice

Cells in vitro or in vivo pre-exposed to low-dose radiation (LDR) or low concentrations of chemical mutagens became more resistant to large-dose radiation-induced DNA or chromosome damage. This was known as radio-adaptive response, for which the exact mechanism was unclear. However, multiple cellular and molecular responses to LDR have been documented, for instance, the induction of some cytokines such as interferon (IFN). Adminstration of exogenous IFN to cultured cells or mice showed marked radio-protection. In the present study, we investigated the in vivo radio-protective effects of extracted human liver RNA (HL-RNA), a known IFN inducer, indirectly to determine the radio-protective action of endogenous IFN. First, mice were administered with 6.25 mg/kg HL-RNA at different times before exposure to radiation and the 24 h pretreatment offered the optimal protective action for HL-RNA on cytogenetic effects in bone marrow cells. When the mice were treated with different concentrations of HL-RNA for 24 h, a wide dose-range (25–100 mg/kg) of HL-RNA resulted in a marked protection from X-ray-induced chromosome aberrations in both bone marrow cells and germ cells. In subsequent experiments, a protective effect of pretreatment with 25 mg/kg HL-RNA for 24 h was also found for radiation-induced micronuclei in polychromatic erythrocytes (PCE), and inhibition of DNA repair ability (unscheduled DNA synthesis, UDS). These results demonstrated that HL-RNA, an IFN inducer, is able to offer significant cytogenetic protection from radiation, implying indirectly that the induction of IFN by LDR may also play a protective role as one of the mechanisms in the induction of the cytogenetic adaptive response.

Molecular Cloning of a Partial cDNA of a Novel Gene in Iron Metabolism

In order to elucidate the possible role played by human hepatic ferritin-associated protein in normal cellular iron metabolism and in iron overload diseases, malignancies and hepatic fibrosis, we attempted to isolate genes for ferritin associated proteins. Here we describe the cloning of a novel cDNA fragment of a 10 kb mRNA transcript from human liver and human T lymphoid (MOLT-4) cells using a short degenerate 5′-primer derived from six amino acid residues of an affinity purified ferritin associated protein from human liver. Northern analysis of this clone has revealed the presence of a 10 kb mRNA species in both human liver and MOLT-4 cell RNA.