Heart tissue contains small and large aggregates of ferritin subunits

Ferritin purified from horse heart and applied to nondenaturing polyacrylamide gel electrophoresis migrated as a single band that stained for both iron and protein. This ferritin contained almost equal amounts of fast- and slow-sedimenting components of 58 S and 3-7 S, which could be separated on sucrose density gradients. Iron removal reduced the sedimentation coefficient of the fast-sedimenting ferritin to 18 S, and sedimentation equilibrium gave a molecular weight 650,000, with some preparations containing ferritin of 500,000 molecular weight as well. Sedimentation rates of the 3 S and 7 S ferritins were not affected by iron removal, and sedimentation equilibrium data were consistent with Mr's 40,000 and 180,000, respectively. Preparations of ferritin extracted from horse spleen contained only 67 S (holo) or 16 S (apo) ferritin and no slow-sedimenting species. When examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, all of the ferritins contained the usual H and L subunits (23 and 20 kDa, respectively), but the slow-sedimenting (3 S and 7 S) heart apoferritins also contained appreciable quantities (ca 25%) of three larger subunits of 42, 55, and 65 kDa. All the subunits reacted positively in Western blots to polyclonal antibodies made against specially purified large heart or spleen ferritins containing only 20- and 23-kDa subunits. Similar results were obtained for ferritins from rat heart. The results indicate that mammalian heart tissue is peculiar not just in having an abnormally large iron-rich ferritin but also in having iron-poor ferritins of much lower molecular weight, partly composed of larger subunits.     

Characterization of myosin heavy chains in cultured aorta smooth muscle cells. A comparative study

Myosin heavy chains (MHCs) from rat aorta smooth muscle cells were analyzed prior to and after these cells were placed into cell culture using sodium dodecyl sulfate-5% polyacrylamide gels, immunoblots, and two-dimensional peptide maps of tryptic digests. Rat aorta smooth muscle cells prior to culture were found to contain two MHCs (mass = 204 and 200 kDa) which cross-reacted with antibodies raised to smooth muscle myosin, but not with antibodies raised to platelet myosin. Tryptic peptide maps of these two MHCs showed no major differences when compared to each other and to maps of vas deferens and uterus smooth muscle MHCs. When rat aorta smooth muscle cells were placed into culture, the MHCs isolated from the cell extracts differed, depending on whether the cells were rapidly growing or postconfluent. Extracts from log-phase cultures contained predominantly MHCs that migrated more rapidly than smooth muscle myosin in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (mass = 196 kDa) and cross-reacted with antibodies raised to platelet myosin, but not to smooth muscle myosin. Tryptic peptide maps of this MHC were very similar to those obtained with MHCs from non-muscle sources such as platelets and fibroblasts. In contrast, extracts from postconfluent rat aorta cell cultures contained three MHCs (mass = 204, 200, and 196 kDa). Using immunoblots and peptide maps, the fastest migrating MHC was found to be identical to the 196-kDa non-muscle MHC, while the two slower migrating MHCs had the same properties as aorta smooth muscle MHCs prior to culture. These results suggest that smooth muscle cells grown in primary culture contain predominantly (greater than 80%) non-muscle myosin while actively growing, but at a postconfluent stage, contain more equivalent amounts of smooth muscle and non-muscle myosins.     

Mouse hepatoma and liver ferritins. Comparative structural studies.

Pure ferritin from male mouse liver produces a single band of monomers (RF = 0.199) with electrophoresis in polyacrylamide gels at pH 9.0. The five sub-bands within this monomeric band appear to represent charge isomers having the same molecular size. Ferritin from BH3 transplantable mouse hepatoma shows two overlapping bands of monomers (RFA = 0.208 and RFB = 0.240); further electrophoretic studies show that these bands represent two subpopulations of molecules differing both in charge and size. Sub-bands are not found in this hepatoma ferritin. The larger tumor ferritin reaches the same end migration position as all liver isoferritins on gradient gels, signifying a very similar or identical molecular size; however, the absence of sub-bands indicates that this hepatoma ferritin differs in charge from the homologous liver proteins. Liver and hepatoma ferritins both produce a single prominent subunit band corresponding to nominal molecular weights of 22 250 and 21 700, with polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and dithiothreitol. With electrophoresis on polyacrylamide gradient slabs containing sodium dodecyl sulfate and dithiothreitol, both liver and hepatoma ferritins now reveal two subunits bands situated at identical positions. The polypeptides of these two closely spaced bands have a nominal molecular weight difference of less than 1000. Neither the hepatoma nor the liver seems to produce the ferritins found in the other tissue. Nevertheless, all these ferritins are composed of the same two types of subunits, albeit in different relative amounts. Observed distinctions in the ferritins from these normal or neoplastic cells must reflect differences in assembly and processing, as well as in the regulated expression of the same ferritin genes.     

Isolation and characterization of chicken muscle ferritin

Two ferritins (fast and slow) have been found to exist in the chicken muscle. Ferritin was isolated from the muscle by means of a method based on pH changes and saline fractioning, followed by purification in Ultrogel AcA-3A and ultracentrifugation at 100,000 g. Identification of the two ferritins shown in the chromatogram was carried out by electrophoresis in polyacrylamide gel, the typical Prussian Blue band with ferrocyamide appearing in both cases. Ferritin characterization was carried out by means of molecular weight determination, amino acid analysis, number of Fe atoms linked by ferritin molecule and other parameters.     

Subpopulations of liver coated vesicles resolved by preparative agarose gel electrophoresis

Rat liver clathrin coated vesicles (CVs) were separated into several distinct subpopulations using non-sieving concentrations of agarose, which allowed the separation of species differing primarily in surface charge. Using preparative agarose electrophoresis (Kedersha, N. L., and L. H. Rome, 1986, Anal. Biochem., in press), the CVs were recovered and analyzed for differences in morphology, coat protein composition, and stripped vesicle protein composition. Coat proteins from different populations appeared identical on SDS PAGE, and triskelions stripped from the different populations showed the same mobility on the agarose gel, suggesting that the mobility differences observed in intact CVs were due to differences in the surface charge of underlying vesicles rather than to variations in their clathrin coats. Several non-coat polypeptides appeared to segregate exclusively with different populations as resolved by two-dimensional electrophoresis. Stripped CVs also exhibited considerable heterogeneity when analyzed by Western blotting: the fast-migrating population was enriched in the mannose 6-phosphate receptor, secretory acetylcholine esterase, and an Mr 195,000 glycoprotein. The slow-migrating population of CVs was enriched in the asialoglycoprotein receptor, and it appeared to contain all detectable concanavalin A-binding polypeptides as well as the bulk of detectable WGA-binding proteins. When CVs were prepared from 125I-asialoorosomucoid-perfused rat liver, ligand was found in the slow-migrating CVs, suggesting that these were endocytic in origin. Morphological differences were also observed: the fast-migrating population was enriched in smaller CVs, whereas the slow-migrating population exhibited an enrichment in larger CVs. As liver consists largely of hepatocytes, these subpopulations appear to originate from the same cell type and probably represent CVs of different intracellular origin and destination.     

Glucocorticoid effects on newly synthesized proteins in muscle and non-muscle cells cultured from neonatal rat hearts

To analyze direct effects of steroids on the rates of synthesis (and/or degradation) of newly synthesized proteins of the rat heart, we have used high resolution two-dimensional gel electrophoresis and autoradiography. A collective steroid domain of nineteen proteins, comprising fifteen with an increased rate of synthesis and four with a decreased rate of synthesis, was consistently seen in cultures of cardiac muscle and non-muscle cells from neonatal rats following 24 h incubation with 10(-7) dexamethasone. Similarly, incubation with 10(-7) M sex steroids, mineralocorticoids, and other glucocorticoids including the highly selective compound RU26988, established the glucocorticoid-specificity of the response. Different subsets of this glucocorticoid domain were seen for collagenase- or trypsin-dispersed primary cultures of cardiac muscle and non-muscle cells or for passaged cultures of cardiac non-muscle cells. Six polypeptides were consistently induced in all cardiac cultures, regardless of cell morphology. Two polypeptides were consistently induced only in those cultures containing cardiac non-muscle cells, whereas protein l, of identical Mr(approximately 52K) and pI (approximately 5.3) to desmin, was induced only in cultures of spontaneously contractile cardiac muscle cells. The glucocorticoid domain proteins described herein represent direct steroid effects on cardiac cells and are therefore candidate mediators of physiological glucocorticoid effects on, for example, differentiation and contractility.     

Size and charge heterogeneity of rat tissue ferritins.

Ferritins purified from horse spleen and from rat liver, kidney, heart and hepatoma were analyzed by quantitative polyacrylamide gel electrophoresis. From the migration characteristics of these ferritins at several gel concentrations, Ferguson plots were constructed and the molecular sizes and charges (apparent valences) together with their statistical variability were obtained by applying Rodbard computer programs to the data. Finally, ellipses were drawn describing the 95% confidence limits of these data for size and charge and were used to identify those ferritins that differed in size and/or charge. By these criteria, many of the tissue ferritins were differentiated from one another in terms of their molecular size and/or charge. Among the various tissue ferritin monomers, the molecular sizes were essentially similar (420 000-490 000) except for the two heart ferritins which were larger (530 000 and 626 000, respectively). However, the estimated charges on rat liver, kidney and hepatoma monomers (30-38 net protons per molecule) differed from that of spleen monomer (51 net protons per molecule) while the larger rat heart ferritin also had a greater charge (83 net protons) than the smaller (40 net protons). Apoferritins prepared chemically by removal of iron from the holoferritins had migration properties indistinguishable from the parent holoferritins. The migration properties of minor (dimeric) ferritin bands on the gels were compared with those of the monomer bands. The molecular sizes of the minor bands were larger than those of the major bands, and were not inconsistent with a doubling in size. However, charge differences varied, being either similar for major and minor forms (spleen ferritin), approximately twice for the minor form (rat hepatoma ferritin) or five times greater for the minor form (rat liver ferritin). These differences in behavior were confirmed by using minimally sieving gels, on which the major bands of horse spleen ferritin failed to separate whereas those of rat liver ferritin were readily separable. It is concluded that dimers of ferritins from different tissues may associate in different ways.     

Isolation of a ferritin from Bacteroides fragilis

A ferritin was isolated from the obligate anaerobe Bacteroides fragilis. Estimated molecular masses were 400 kDa for the holomer and 16.7 kDa for the subunits. A 30-residue N-terminal amino acid sequence was determined and found to resemble the sequences of other ferritins (human H-chain ferritin, 43% identity; Escherichia coli gen-165 product, 37% identity) and to a lesser degree, bacterioferritins (E. coli bacterioferritin, 20% identity). The protein stained positively for iron, and incorporated 59Fe when B. fragilis was grown in the presence of [59Fe]citrate. However, the isolated protein contained only about three iron atoms per molecule, and contained no detectable haem. This represents the first isolation of a ferritin protein from bacteria. It may alleviate iron toxicity in the presence of oxygen.     

A biochemical comparison of normal human liver and hepatocellular carcinoma ferritins.

1. The iron contents, gel migration rates and isoelectric-focusing patterns of normal liver and hepatocellular carcinoma ferritins from the same patients were compared. 2. Sucrose-density-gradient centrifugation showed that the number of iron atoms per ferritin molecule was decreased to approximately half in carcinoma tissue when compared with normal liver. 3. On electrophoresis, hepatocellular carcinoma ferritin migrates faster and is therefore more negatively charged than normal liver ferritin, thus refuting the general view that the more negatively charged a ferritin molecule the greater its iron content. 4. Comparison of tumour and normal liver ferritin subunit compositions on acid/urea/polyacrylamide gels showed hepatocellular carcinoma ferritin to contain an additional, more negatively charged, subunit to normal liver ferritin. 5. Isoelectric focusing showed that hepatocellular carcinoma tissue contains isoferritins with isoelectric points intermediate between the ranges of normal liver and normal heart isoferritins.     

Isolation, purification and characterization of bovine spleen ferritin

Ferritin was isolated from bovine spleen and used to prepare apoferritin and reconstituted ferritin. The mol. wt of bovine ferritin was 464,000 with monomer subunits about 18,000-19,500. Gel electrophoresis showed three bands each for ferritin, apoferritin and reconstituted ferritin; all stained for protein and carbohydrate. Only apoferritin failed to stain for iron. Bovine ferritin had higher concentrations of proline, threonine, and valine than equine or human ferritin. The iron:protein ratio of bovine ferritin was 0.161 and of equine ferritin was 0.192. After iron uptake by the apoferritins the iron:protein ratios were 0.186 and 0.278 for the bovine and equine ferritins, respectively.     

Molecular cloning,expression and isolation of ferritins from two tick species--Ornithodoros moubata and Ixodes ricinus

Genes encoding ferritins were isolated and cloned from cDNA libraries of hard tick Ixodes ricinus and soft tick Ornithodoros moubata. Both tick ferritins are composed of 172 amino-acid residues and their calculated mass is 19,667.2 Da and 19,974.5 Da for I. ricinus and O. moubata, respectively. The sequences of both proteins are closely related to each other as well as to the ferritin from another tick species Dermacentor variabilis (>84% similarity). The proteins contain the conserved motifs for ferroxidase center typical for heavy chains of vertebrate ferritins. The stem-loop structure of a putative iron responsive element was found in the 5' untranslated region of ferritin mRNA of both ticks. Antibodies against fusion ferritin from O. moubata were raised in a rabbit and used to monitor the purification of a small amount of ferritins from both tick species. The authenticity of ferritin purified from O. moubata was confirmed by mass-fingerprinting analysis. In the native state, the tick ferritins are apparently larger (~500 kDa) than horse spleen ferritin (440 kDa). On SDS-PAGE tick ferritins migrate as a single band of about 21 kDa. These results suggest that tick ferritins are homo-oligomers of 24 identical subunits of heavy-chain type. The Northern blot analysis revealed that O. moubata ferritin mRNA level is likely not up-regulated after ingestion of a blood meal.     

Correlation of myosin isoforms with anatomical divisions in equine musculus biceps brachii.

The biceps brachii of horses is subdivided into a lateral and medial head. Electrophoresis of samples from the lateral head revealed three slow-migrating native myosin isoforms, including one that does not correspond to slow myosin isoforms described for other mammalian muscles. In contrast, the medial head contained a single slow isoform. Both the lateral and medial heads contained three fast-migrating isoforms corresponding with the FM-2, FM-3 and FM-4 isoforms reported for other mammalian fast-twitch muscle fibers. Electrophoresis of myosin heavy chains (MHCs) revealed only two MHC bands, one fast-migrating band that comigrates with rat type I MHC and a second slower-migrating band that comigrates with rat type IIa MHC. Quantitation of the histochemical data is correlated with densitometric analysis of MHCs in the medial and lateral heads of biceps brachii and is consistent with previously hypothesized functional specializations of this muscle.     

Comparison of cytosolic products formed in rat liver in response to parenteral and dietary iron loading

Two different methods were used to create a situation of iron (Fe) overload in rats. One group of rats received Fe dextran, and another group of rats received a carbonyl Fe-enriched diet. The ferritins present in the liver cytosol of these rats were isolated and compared. From each group, two cytosolic products were isolated with the use of ultracentrifugation: a cytosolic ferritin fraction (CF) and a (slower sedimenting) light ferritin fraction (CLF). There were no differences with respect to the protein coat (subunit composition and amino acid analysis). Analysis of the Fe core revealed that the two CF fractions were similar, whereas the two CLF fractions differed with respect to their Fe content and to the packing of their cores. The carbonyl CLF product contained less Fe atoms/molecule, which, moreover, seemed to be packed in a less compact way.     

Characteristics of structure, composition, mass spectra, and iron release from the ferritin of shark liver (Sphyrna zygaena)

The ferritin consists of a protein shell constructed of 24 subunits and an iron core. The liver ferritin of Sphyrna zygaena (SZLF) purified by column chromatography is a protein composed of eight ferritins containing varying iron numbers ranging from 400+/-20 Fe3+/SZLF to 1890+/-20 Fe3+/SZLF within the protein shell. Nature SZLF (SZLFN) consisting of holoSZLF and SZLF with unsaturated iron (SZLFUI) to have been purified with polyacrylamide gel electrophoresis (PAGE) exhibited five ferritin bands with different pI values ranging from 4.0 to 7.0 in the gel slab of isoelectric focusing (IEF). HoloSZLF purified by PAGE (SZLFE) not only had 1890+/-20 Fe3+/SZLFE but also showed an identical size of iron core observed by transmission electron microscopy (TEM). Molecular weight of approximately 21 kDa for SZLFE subunit was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Four peaks of molecular ions at mass/charge (m/z) ratios of 10611.07, 21066.52, 41993.16, and 63555.64 that come from the SZLFE were determined by matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF MS), which were identified as molecular ions of the ferritin subunit (M+) and its polymers, namely, [M]2+, [M]+, [2M]+, and [3M]+, respectively. Both SZLFE and a crude extract from shark liver of S. zygaena showed similar kinetic characteristics of complete iron release with biphasic behavior. In addition, a combined technique of visible spectrometry and column chromatography was used for studying ratio of phosphate to Fe3+ within the SZLFE core. Interestingly, this ratio maintained invariable even after the iron release, which differed from that of other mammal ferritins.     

Molecular size heterogeneity of ferritin in mouse liver

As much as 4% of the total protein in pure liver ferritin from mice with short-term parenteral iron overload produces a minor band migrating anodally to the major (alpha) band of holoferritin with non-denaturing polyacrylamide gel electrophoresis. The components in this minor band and the alpha band have been isolated to purity by preparative electrophoretic fractionation. The protein in the minor band is ferritin, since it contains ferric iron and fulfills defining criteria at the level of biochemistry, immunology and ultrastructure. Native polyacrylamide electrophoresis with pore-size-gradient gels shows that the ferritin molecules in the minor band have a slightly smaller diameter than the holoferritin in the alpha band. Isoelectric focusing reveals that the smaller ferritin has an identical number and range of charge isomers (pI 4.9-5.3) as the larger ferritin, but the relative amount of each size class within some isoferritin bands differs. The smaller ferritin molecules are structurally intact and are made from polypeptide subunits with Mr 18 000; the larger ferritin molecules have subunits with Mr 22 000. The minor species of hepatic ferritin thus has a smaller molecular size because it is made mainly from smaller subunits. No minor electrophoretic band can be detected in liver ferritin obtained from mice with normal iron levels. These results demonstrate that siderosis induces the formation of molecular size polymorphism (macroheterogeneity) in mouse liver ferritin. The new smaller hepatic ferritin could serve to redistribute excess iron into the main storage organs during the early response to iron overload, since it appears to be identical to one of the two types of serum ferritin molecules present in these siderotic mice.     

Characterization of ferritin from human placenta. Implications for analysis of tissue specificity and microheterogeneity of ferritins

Mammalian ferritins can be resolved into multiple components by isoelectric focusing, and each tissue contains a characteristic subset of isoferritins. Ferritin isolated from human liver was compared to acidic ferritin isolated from mid-gestational human placenta to define a structural basis for ferritin heterogeneity. Placenta ferritin contained several major bands with isoelectric points in the range of pI = 4.7-5.0 which were more acidic than the predominant isoferritins of human liver. Ferritin from each tissue was resistant to denaturation by 10 M urea and appeared to be identical by electron microscopy. Circular dichroism measurements revealed that placenta ferritin had substantially less ordered secondary structure than liver ferritin. Both types of ferritin contained only two subunits when analyzed by electrophoresis in sodium dodecyl sulfate gels, but isoelectric focusing of dissociated subunits in urea revealed 6-7 different components. In this system, placenta ferritin was enriched in the more acidic subunits and it completely lacked the most basic subunits noted in liver ferritin; placental ferritin had no unique components. Differences in isoelectric points among assembled ferritins from these two tissues appear to result from different proportions of these acidic and basic subunits.     

Macrophage interaction of HDL subclasses separated by free flow isotachophoresis

Preparative isotachophoresis (ITP) was used for the fractionation of fasting and postprandial high density lipoproteins (HDL) according to their net charge in the absence of molecular sieve effects. Three major HDL subpopulations with fast, intermediate, and slow mobility have been recognized. Particle size analysis by gradient gel electrophoresis has shown that in the fast-migrating subpopulation particles dominate with a size of HDL3a and HDL2b. The subpopulation with intermediate mobility contains particles with a size between HDL2a and HDL3b, while in the slow migrating subpopulation particles dominate with a size of HDL2b, HDL3a, and HDL3c. The fast-migrating subpopulation is rich in apoA-I and phosphatidylcholine. The particles of this fraction bind at 4 degrees C to HDL receptors on macrophages with high affinity (KD = 7.71 micrograms/ml; Bmax = 245.6 ng). The subpopulations with intermediate mobility is rich in apoA-II, apoE, C apolipoproteins, cholesteryl esters, and sphingomyelin. Its affinity to HDL receptors (KD = 17.7 micrograms/ml; Bmax = 198.4 ng) is lower than that of the HDL particles in the fast-migrating subfraction. The slow-migrating subpopulation consists of particles rich in apoA-IV and is associated with a high LCAT activity. This fraction expresses the highest nonspecific binding to mouse peritoneal macrophages compared to the other HDL fractions and contains only a small amount of particles that interact with HDL receptors by high affinity binding (KD = 7.3 micrograms/ml; Bmax = 95.9 ng). In 37 degrees C binding experiments the fast-migrating subfraction reveals the highest total cell-associated activity. 72% of which is trypsin-resistant. The other subfractions express a lower total cell-associated activity and 45% of the activity of the intermediate- and 43% of the activity of the slow-migrating fraction is trypsin-sensitive. When the HDL fractions are isolated from postprandial sera of the same donor, the fast-migrating particles bind at 4 degrees C with a higher affinity (KD = 4.6 micrograms/ml) while no significant changes are observed in the intermediate- and slow-migrating subpopulations. The slow- and the fast-migrating HDL subpopulations isolated from fasting serum have a high capacity to promote cholesterol removal from macrophages. We hypothesize that the HDL subpopulations rich in apoA-I promote cholesterol removal predominantly via the interaction with HDL receptors, while apoA-IV-rich HDL particles receive their driving force for cholesterol efflux from the concomitant action of LCAT via a predominantly nonspecific interaction of the particles with the cell surface.(ABSTRACT TRUNCATED AT 400 WORDS)     

Homologous terminal sequences of the genome double-stranded RNAs of bluetongue virus.

The 3'-terminal sequences of the 10 double-stranded RNA genome segments of bluetongue virus (serotypes 10 and 11) were determined. The double-stranded RNAs were 3' labeled with [5'-32P]pCp and resolved into 10 segments by electrophoresis. After denaturation, the two complementary strands of segments 4 through 10 were resolved into fast- and slow-migrating species by polyacrylamide gel electrophoresis, and their 3' end sequences were determined. Complete RNase T1 digestion of the individual 3'-labeled double-stranded RNA segments yielded two labeled oligonucleotides, one of which migrated faster than the other on 20% polyacrylamide-7 M urea gels. Sequence analyses of the two oligonucleotides of segments 4 through 10 confirmed the corresponding RNA sequence data. For RNA segments 1 through 3 the oligonucleotide analyses gave comparable results. The 3'-terminal sequences of the fast-migrating RNA species were HOCAAUUU. . . ; those of the slow-migrating RNA species were HOCAUUCACA. . . . Similar results were obtained for double-stranded RNA from bluetongue virus serotypes 10 and 11. Beyond the common termini, the sequences for each segment varied considerably.     

Biochemical and morphological differences between fibroblasts and myoblasts from embryonic chicken skeletal muscle

Summary Non-myogenic cells were isolated from the breast muscle of 10-day-old chicken embryos employing Percoll density centrifugation. In culture, these cells exhibited the spread out, stellate morphology of fibroblast-like cells. They also exhibited receptor-mediated binding of plateletderived growth factor (PDGF). Such binding was not detected in cultures of predominantly myogenic cells isolated by the Percoll density centrifugation from the same muscle. Percoll-isolated myogenic and fibrogenic cell populations were also analyzed by two-dimensional polyacrylamide gel electrophoresis immediately after removal from the muscle. This analysis revealed at least six polypeptides specific to the fibroblasts but not detected in the myogenic cell population. In addition, at least eight polypeptides found in the myogenic population were barely detectable, or lacking altogether from the fibroblast-like cells. Ultrastructural analysis of the freshly isolated cells demonstrated that the fibroblasts were larger than the myoblasts and that their cytoplasm contained many vesicles. We conclude that the fibrogenic and myogenic cells isolated by Percoll from embryonic muscle express cell type-specific characteristics. Moreover, based on the PDGF binding studies, the fibrogenic cells can be categorized as true fibroblasts.