IFN-γ engenders strong anti-proliferative responses, in part through activation of p53. However, the long-known IFN-γ-dependent upregulation of human Trp-tRNA synthetase (TrpRS), a cytoplasmic enzyme that activates tryptophan to form Trp-AMP in the first step of protein synthesis, is unexplained. Here we report a nuclear complex of TrpRS with the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) and with poly (ADP-ribose) polymerase 1 (PARP-1), the major PARP in human cells. The IFN-γ-dependent poly (ADP-ribosyl)ation of DNA-PKcs (which activates its kinase function) and concomitant activation of p53 were specifically prevented by Trp-SA, an analog of Trp-AMP that disrupted the TrpRS/DNA-PKcs/PARP-1 complex. The connection of TrpRS to p53 signaling in vivo was confirmed in a vertebrate system. These and further results suggest a surprising evolutionary expansion of the protein synthesis apparatus to a nuclear role that links major signaling pathways.
Aminoacyl-tRNA synthetases (AARSs) catalyze aminoacylation of tRNAs in the cytoplasm. Surprisingly, AARSs also have critical extracellular and nuclear functions. Evolutionary pressure for new functions might be manifested by splice variants that skip only an internal catalytic domain (CD) and link non-catalytic N- and C-terminal polypeptides. Using disease-associated histidyl-tRNA synthetase (HisRS) as an example, we discovered an expressed 171 amino acid protein (HisRSΔCD) that deleted the entire CD, and joined an N-terminal WHEP to the C-terminal anticodon-binding domain (ABD). X-ray crystallographic and 3-D NMR methods revealed the first structures of human HisRS and HisRSΔCD. In contrast to homodimeric HisRS, HisRSΔCD is monomeric, where rupture of the ABD’s packing with CD resulted in a dumbbell-like structure of flexibly linked WHEP and ABD domains. In addition, the ABD of HisRSΔCD presents a new local conformation. This natural internally deleted HisRS suggests evolutionary pressure to reshape AARS tertiary and quaternary structures for repurposing.
Human respiratory syncytial virus (RSV) is a serious pediatric pathogen of the lower respiratory tract. Currently, there is no clinically approved vaccine against RSV infection. Recent studies have shown that helper-dependent adenoviral (HDAd) vectors may represent effective and safe vaccine vectors. However, viral challenge has not been investigated following mucosal vaccination with HDAd vector vaccines.
To explore the role played by HDAd as an intranasally administered RSV vaccine vector, we constructed a HDAd vector encoding the codon optimized fusion glycoprotein (Fsyn) of RSV, designated HDAd-Fsyn, and delivered intranasally HDAd-Fsyn to mice.
RSV-specific humoral and cellular immune responses were generated in BALB/c mice, and serum IgG with neutralizing activity was significantly elevated after a homologous boost with intranasal (i.n.) application of HDAd-Fsyn. Humoral immune responses could be measured even 14 weeks after a single immunization. Immunization with i.n. HDAd-Fsyn led to effective protection against RSV infection on challenge.
The results indicate that HDAd-Fsyn can induce powerful systemic immunity against subsequent i.n. RSV challenge in a mouse model and is a promising candidate vaccine against RSV infection.
Human respiratory syncytial virus; Helper-dependent adenovirus vectors; Fusion glycoprotein; Protective immunity; Immune responses
Radioiodine therapy has proven to be a safe and effective approach in the treatment of differentiated thyroid cancer. Similar treatment strategies have been exploited in nonthyroidal malignancies by transfecting hNIS gene into tumor cells or xenografts. However, rapid radioiodine efflux is often observed after radioiodine uptake, limiting the overall antitumor effects. In this study, we aimed at constructing multicistronic co-expression of hNIS and hTPO genes in tumor cells to enhance the radioiodine uptake and prolong the radioiodine retention. Driven by the cytomegalovirus promoter, hNIS and hTPO were simultaneously inserted into the expression cassette of adenoviral vector. An Ad5 viral vector (Ad-CMV-hTPO-T2A-hNIS) was assembled as a gene therapy vehicle by Gateway technology and 2A method. The co-expression of hNIS and hTPO genes was confirmed by a double-label immunofluorescence assay. The radioiodine (125I) uptake and efflux effects induced by co-expression of hNIS and hTPO genes were determined in transfected and non-transfected PC-3 cells. Significantly higher uptake (6.58 ± 0.56 fold, at 1 h post-incubation) and prolonged retention (5.47 ± 0.36 fold, at 1 h of cell efflux) of radioiodine (125I) were observed in hNIS and hTPO co-expressed PC-3 cells as compared to non-transfected PC-3 cells. We concluded that the new virus vector displayed favorable radioiodine uptake and retention properties in hNIS-hTPO transfected PC-3 cells. Our study will provide valuable information on improving the efficacy of hNIS-hTPO co-mediated radioiodine gene therapy.
Gene therapy; prostate cance; hNIS; hTPO; gateway cloning system
AIM: To study the effect of H2 gas on liver injury in massive hepatectomy using the Intermittent Pringle maneuver in swine.
METHODS: Male Bama pigs (n = 14) treated with ketamine hydrochloride and Sumianxin II as induction drugs followed by inhalation anesthesia with 2% isoflurane, underwent 70% hepatotectomy with loss of bleeding less than 50 mL, and with hepatic pedicle occlusion for 20 min, were divided into two groups: Hydrogen-group (n = 7), the pigs with inhalation of 2% hydrogen by the tracheal intubation during major hepatotectomy; Contrast-group (n = 7), underwent 70% hepatotectomy without inhalation of hydrogen. Hemodynamic changes and plasma concentrations of alanine aminotransferase (ALT), aspartate aminotransferase (AST), hyaluronic acid (HA), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and malondialdehyde (MDA) in liver tissue were measured at pre-operation, post-hepatotectomy (PH) 1 h and 3 h. The apoptosis and proliferating cell nuclear antigen (PCNA) expression in liver remnant were evaluated at PH 3 h. Then we compared the two groups by these marks to evaluate the effect of the hydrogen in the liver injury during major hepatotectomy with the Pringle Maneuver in the swine.
RESULTS: There were no significant differences in body weight, blood loss and removal liver weight between the two groups. There was no significant difference in changes of portal vein pressure between two groups at pre-operation, PH 30 min, but in hydrogen gas treated-group it slightly decrease and lower than its in Contrast-group at PH 3 h, although there were no significant difference (P = 0.655). ALT and AST in Hydrogen-group was significantly lower comparing to Contrast-group (P = 0.036, P = 0.011, vs P = 0.032, P = 0.013) at PH 1 h and 3 h, although the two groups all increased. The MDA level increased between the two group at PH 1 h and 3 h. In the hydrogen gas treated-group, the MDA level was not significantly significant at pre-operation and significantly low at PH 1 h and 3 h comparing to Contrast-group (P = 0.0005, P = 0.0004). In Hydrogen-group, the HA level was also significantly low to Contrast-group (P = 0.0005, P = 0.0005) although the two groups all increased at PH 1 h and 3 h. The expression of cluster of differentiation molecule 31 molecules Hydrogen-group was low to Contrast-group. However, PCNA index (%) was not statistically significant between the two groups (P = 0.802). Microphotometric evaluation of apoptotic index (AI) in terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-stained tissue after hepatotectomy for 3h, the AI% level in the hydrogen was significantly low to Contrast-group (P = 0.012). There were no significant difference between Hydrogen-group and Contrast-group at pre-operation (P = 0.653, P = 0.423), but after massive hepatotectomy, the TNF-α and IL-6 levels increase, and its in Hydrogen-group was significantly low compared with Contrast-group (P = 0.022, P = 0.013, vs P = 0.016, P = 0.012), respectively. Hydrogen-gas inhalation reduce levels of these markers and relieved morphological liver injury and apoptosis.
CONCLUSION: H2 gas attenuates markedly ischemia and portal hyperperfusion injury in pigs with massive hepatotectomy, possibly by the reduction of inflammation and oxidative stress, maybe a potential agent for treatment in clinic.
Massive hepatotectomy; Hydrogen gas; Anti-oxidant; Hyperperfusion; Malondialdehyde; Oxidative stress
Emerging mechanism for tRNA fragment-based translational regulation expands our understanding of the non-decoding roles of tRNA.
In the cation of the title compound, [Ru(C15H10ClN3)2]Cl2·2H2O, the metal atom exhibits a distorted octahedral coordination geometry provided by the N atoms of two tridentate terpyridine ligands. The ligands are approximately planar [maximum deviation = 0.156 (5) Å] and form a dihedral angle of 87.0 (3)°. In the crystal, the cations, anions and water molecules are linked into a three-dimensional network by C—H⋯Cl, C—H⋯O and O—H⋯Cl hydrogen bonds.
Breast cancer is the most common cancer and the leading cause of cancer mortality in women worldwide. Hypoxia is an important factor involved in the progression of solid tumors and has been associated with various indicators of tumor metabolism, angiogenesis and metastasis. But little is known about the contribution of Hypoxia-Inducible Factor-2a (HIF-2a) to the drug resistance and the clinicopathological characteristics in breast cancer.
Immunohistochemistry was employed on the tissue microarray paraffin sections of surgically removed samples from 196 invasive breast cancer patients with clinicopathological data. The correlations between the expression of HIF-2a and ABCG2 as well as other patients' clinicopathological data were investigated.
The results showed that HIF-2a was expressed in different intensities and distributions in the tumor cells of the breast invasive ductal carcinoma. A positive staining for HIF-2a was defined as a brown staining observed mainly in the nucleus. A statistically significant correlation was demonstrated between HIF-2a expression and ABCG2 expression (p = 0.001), histology-grade (p = 0.029), and Ki67 (p = 0. 043) respectively.
HIF-2a was correlated with ABCG2 expression, histology-grade and Ki67 expression in breast invasive ductal carcinoma. HIF-2a could regulate ABCG2 in breast cancer cells, and could be a novel potential bio-marker to predict chemotherapy effectiveness. The hypoxia/HIF-2a/ABCG2 pathway could be a new mechanism of breast cancer multidrug-resistance.
HIF-2a; ABCG2(BCRP); Histology-grade; Ki67; Breast invasive ductal cancer; IHC; Tissue microarray; MDR
Objective: To investigate the expression of glioma-associated oncogene homolog 1(Gli-1) in colon cancer and its association with clinicopathological parameters and postoperative liver metastasis.
Methods: Expression of Gli-1 was detected by immunohistochemistry in paraffin-embedded specimens of 96 cases of colon cancer. Relationship between Gli-1 expression and clinicopathological parameters, postoperative liver metastasis were analyzed.
Results: Gli-1 protein expression was significantly increased in colon cancer tissues compared to normal colon tissues (P = 0.037). Gli-1 expression in colon tissues was increased in patients with lymph node metastases (P = 0.022) and higher T stages (P = 0.030). Postoperative live metastasis-free survival period was significantly longer in low Gli-1 expression group than that of high Gli-1 expression group (48.22±10.03 months vs 20.46±6.32 months, P=0.001). Multivariate analysis showed that Gli-1 expression level is an independent prognostic factor for postoperative live metastasis-free survival.
Conclusion: Colon cancer is associated with an upregulation of Gli-1 protein expression in colon tissues. In patients with colon cancer, Gli-1 expression level is closely related to lymph node metastases, T stages and postoperative live metastasis-free survival periods, indicative of a possible role of Gli-1 expression in colon cancer progression.
Colon neoplasm; Glioma-associated oncogene 1; Liver metastasis.
Aminoacyl tRNA synthetases, components of the translation apparatus, have alternative functions outside of translation. The structural and mechanistic basis of these alternative functions is of great interest. As an example, reverse transcription of the HIV genome is primed by a human lysine-specific tRNA (tRNALys3) that is packaged (into the virion) by the HIV gag protein with lysyl-tRNA synthetase (LysRS). Not understood is the structural basis for simultaneous packaging of tRNALys3, LysRS and Gag. Here ab initio computational methods, together with our recent high resolution 3-D structure of human LysRS, produced an energy-minimized model where Gag, tRNALys, and LysRS form a ternary complex. Interestingly, the model requires normally homodimeric LysRS to dissociate into a monomer that bridges between Gag and tRNALys3. Earlier experiments of others, and new experiments presented here, which tested an engineered dissociated form of LysRS, were consistent with the ab initio ‘bridging -monomer’ model. The results support an emerging theme that alterative functions of tRNA synthetases may come in part from protein surfaces exposed by dynamic equilibria.
Charcot-Marie-Tooth disease type 2D (CMT2D) is a dominantly inherited peripheral neuropathy caused by missense mutations in the glycyl-tRNA synthetase gene (GARS). In addition to GARS, mutations in three other tRNA synthetase genes cause similar neuropathies, although the underlying mechanisms are not fully understood. To address this, we generated transgenic mice that ubiquitously over-express wild-type GARS and crossed them to two dominant mouse models of CMT2D to distinguish loss-of-function and gain-of-function mechanisms. Over-expression of wild-type GARS does not improve the neuropathy phenotype in heterozygous Gars mutant mice, as determined by histological, functional, and behavioral tests. Transgenic GARS is able to rescue a pathological point mutation as a homozygote or in complementation tests with a Gars null allele, demonstrating the functionality of the transgene and revealing a recessive loss-of-function component of the point mutation. Missense mutations as transgene-rescued homozygotes or compound heterozygotes have a more severe neuropathy than heterozygotes, indicating that increased dosage of the disease-causing alleles results in a more severe neurological phenotype, even in the presence of a wild-type transgene. We conclude that, although missense mutations of Gars may cause some loss of function, the dominant neuropathy phenotype observed in mice is caused by a dose-dependent gain of function that is not mitigated by over-expression of functional wild-type protein.
Mutations in the glycyl-tRNA synthetase gene (GARS) cause Charcot-Marie-Tooth disease type 2D, a disease characterized by neuronal axon loss in the arms and legs, resulting in weakness and sensory problems. The GARS protein is essential for protein synthesis in every cell, and it has been difficult to determine whether the mutations result in disease because they impair this function or whether GARS somehow becomes toxic when it is mutated. We generated mice that overexpress normal GARS and mated these to two different mouse models of the disease to determine whether a restoration of normal function could prevent the disease. These crosses demonstrated that the mutant forms of GARS are toxic, and this toxic effect increases as the amount of mutant protein increases. Furthermore, this toxicity cannot be reduced or prevented by providing additional normal GARS. Therefore, these results suggest that, for most patients, therapies need to specifically target the mutant form of GARS or the toxic function.
To investigate the association between neck circumference and central obesity, overweight, and metabolic syndrome in Chinese individuals with type 2 diabetes.
RESEARCH DESIGN AND METHODS
A total of 3,182 diabetic subjects (aged 20–80 years) were recruited from 15 community health centers in Beijing using a multistage random sampling approach.
Receiver operating characteristic analysis showed that the area under the curve for neck circumference and central obesity was 0.77 for men and 0.75 for women (P < 0.001). Furthermore, a neck circumference of ≥38 cm for men and ≥35 cm for women was the best cutoff point for determining overweight subjects. A neck circumference of ≥39 cm for men and ≥35 cm for women was the best cutoff point to determine subjects with metabolic syndrome.
In the present study, neck circumference is positively related with BMI, waist circumference, and metabolic syndrome in Chinese individuals with type 2 diabetes.
ABCG2 is an ABC transporter. It has been demonstrated that endogenous ABCG2 expression in certain cancers is a possible reflection of the differentiated phenotype of the cell of origin and likely contributes to intrinsic drug resistance. But little is known about the contribution of ABCG2 to the drug resistance and the clinicopathological characteristics in breast cancer. In the present study, we investigated the expression of ABCG2 and the correlations between ABCG2 expression and patients' clinicopathological and biological characteristics.
Immunohistochemistry was employed on the tissue microarray paraffin sections of surgically removed samples from 196 breast cancer patients with clinicopathological data.
The results showed that ABCG2 was expressed in different intensities and distributions in the tumor cells of the breast invasive ductal carcinoma. A positive stain for ABCG2 was defined as a brown stain observed in the cytoplasm and cytomembrane. A statistically significant correlation was demonstrated between ABCG2 expression and HER-2 expression (p = 0.001), lymph node metastasis (p = 0.049), and clinical stage (p = 0.015) respectively.
ABCG2 correlated with Her-2 expression, lymph node metastasis and clinical stage in breast invasive ductal carcinoma. It could be a novel potential bio-marker which can predict biological behavior, clinical progression, prognosis and chemotherapy effectiveness.
ABCG2(BCRP); Her-2; Lymph node metastasis; Clinical stage; Correlation; Breast invasive ductal cancer; Immunohistochemistry; Tissue microarray
Over the course of evolution, eukaryote aminoacyl tRNA synthetases progressively added domains and motifs that have no essential connection to aminoacylation reactions. Their accretive addition to virtually all tRNA synthetase correlates with the progressive evolution and complexity of eukaryotes. Based on recent experimental findings focused on a few of these additions, and analysis of the tRNA synthetase proteome, we propose that these additions are markers for synthetase-associated functions beyond translation.
In the title compound, C19H16N2O2, the benzene ring and naphthyl ring system are inclined at a dihedral angle of 16.1 (3)°. An intramolecular O—H⋯N hydrogen bond influences the molecular conformation. In the crystal, molecules are linked through N—H⋯O hydrogen bonds into chains running along the a axis.
In the title compound, C19H16N2O2, the benzene ring and the naphthyl ring system form a dihedral angle of 8.7 (3)° and an intramolecular O—H⋯N hydrogen bond generates an S(6) ring. In the crystal, molecules are linked by N—H⋯O hydrogen bonds, forming C(4) chains propagating in .
E. coli alanyl-tRNA synthetase is recalcitrant to crystallization. A group of leucine substitutions has transformed the protein.
Although Escherichia coli alanyl-tRNA synthetase was among the first tRNA synthetases to be sequenced and extensively studied by functional analysis, it has proved to be recalcitrant to crystallization. This challenge remained even for crystallization of the catalytic fragment. By mutationally introducing three stacked leucines onto the solvent-exposed side of an α-helix, an engineered catalytic fragment of the synthetase was obtained that yielded multiple high-quality crystals and cocrystals with different ligands. The engineered α-helix did not form a leucine zipper that interlocked with the same α-helix from another molecule. Instead, using the created hydrophobic spine, it interacted with other surfaces of the protein as a leucine half-zipper (LHZ) to enhance the crystal lattice interactions. The LHZ made crystal lattice contacts in all crystals of different space groups. These results illustrate the power of introducing an LHZ into helices to facilitate crystallization. The authors propose that the method can be unified with surface-entropy reduction and can be broadly used for protein-surface optimization in crystallization.
surface engineering; surface-entropy reduction; leucine zippers; aminoacyl-tRNA synthetases
Protein multi-functionality is an emerging explanation for the complexity of higher organisms. In this regard, while aminoacyl tRNA synthetases catalyze amino acid activation for protein synthesis, some also act in pathways for inflammation, angiogenesis, and apoptosis. How multiple functions evolved and their relationship to the active site is not clear. Here structural modeling analysis, mutagenesis, and cell-based functional studies show that the potent angiostatic, natural fragment of human TrpRS associates via Trp side chains that protrude from the cognate cellular receptor VE-cadherin. Modeling indicates that (I prefer the way it was because the conclusion was reached not only by modeling, but more so by experimental studies.)VE-cadherin Trp side chains fit into the Trp-specific active site of the synthetase. Thus, specific side chains of the receptor mimic (?) amino acid substrates and expand the functionality of the active site of the synthetase. We propose that orthogonal use of the same active site may be a general way to develop multi-functionality of human tRNA synthetases and other proteins.
Known as an essential component of the translational apparatus, the aminoacyl-tRNA synthetase family catalyzes the first step reaction in protein synthesis, that is, to specifically attach each amino acid to its cognate tRNA. While preserving this essential role, tRNA synthetases developed other roles during evolution. Human tRNA synthetases, in particular, have diverse functions in different pathways involving angiogenesis, inflammation and apoptosis. The functional diversity is further illustrated in the association with various diseases through genetic mutations that do not affect aminoacylation or protein synthesis. Here we review the accumulated knowledge on how human tRNA synthetases used structural inventions to achieve functional expansions.
Aminoacyl-tRNA synthetase; non-canonical function; human; structure; multisynthetase complex; disease
The crystal structure of tryptophanyl-tRNA synthetase from T. maritima unexpectedly revealed an iron–sulfur cluster bound to the tRNA anticodon-binding region.
A novel aminoacyl-tRNA synthetase that contains an iron–sulfur cluster in the tRNA anticodon-binding region and efficiently charges tRNA with tryptophan has been found in Thermotoga maritima. The crystal structure of TmTrpRS (tryptophanyl-tRNA synthetase; TrpRS; EC 22.214.171.124) reveals an iron–sulfur [4Fe–4S] cluster bound to the tRNA anticodon-binding (TAB) domain and an l-tryptophan ligand in the active site. None of the other T. maritima aminoacyl-tRNA synthetases (AARSs) contain this [4Fe–4S] cluster-binding motif (C-x
2-C). It is speculated that the iron–sulfur cluster contributes to the stability of TmTrpRS and could play a role in the recognition of the anticodon.
TM0492; tryptophanyl-tRNA ligase; tryptophanyl-tRNA synthetase class I; iron–sulfur clusters; structural genomics
Mistranslation from confusion of serine for alanine by alanyl-tRNA synthetases (AlaRSs) has profound functional consequences1-3. Throughout evolution, two editing-checkpoints prevent disease-causing mistranslation from confusing glycine or serine for alanine at the active site of AlaRS. In both bacteria and mice, Ser poses a bigger challenge than Gly1,2. One checkpoint is the AlaRS editing center, while the other is from widely distributed AlaXps—free-standing, genome-encoded editing proteins that clear Ser-tRNAAla. The paradox of misincorporating both a smaller (glycine) and a larger amino acid (serine) suggests a deep conflict for nature-designed AlaRS. To understand the chemical basis for this conflict, kinetic and mutational analysis, together with nine crystal structures, provided snapshots of adenylate formation for each amino acid. An inherent dilemma is posed by constraints of a structural design that pins down the α–amino group of the bound amino acid using an acidic residue. This design, of more than 3 billion years, creates a serendipitous interaction with the serine OH that is difficult to avoid. Apparently not able to find better architecture for recognition of alanine, the serine misactivation problem was solved through free-standing AlaXps, which appeared contemporaneously with early AlaRSs. The results reveal unconventional problems and solutions arising from the historical design of the protein synthesis machinery.
Aminoacyl-tRNA synthetases are known for catalysis of aminoacylation. Significantly, some mammalian synthetases developed cytokine functions possibly linked to disease-causing mutations in tRNA synthetases. Not understood is how epitopes for cytokine signaling were introduced into catalytic scaffolds without disturbing aminoacylation. Here we investigate human tyrosyl-tRNA synthetase, where a catalytic-domain surface helix—next to the active site—was recruited for IL-8-like cytokine signaling. Taking advantage of our high-resolution structure, the reciprocal impact of rational mutations designed to disrupt aminoacylation or cytokine signaling was investigated with multiple assays. The collective analysis demonstrated a protective fine–structure separation of aminoacylation from cytokine activities within the conserved catalytic domain. As a consequence, disease-causing mutations affecting cell signaling can arise without disturbing aminoacylation. These results with TyrRS also predict the previously unknown binding conformation of IL-8-like CXC cytokines.
Disease-causing mutations occur in genes for aminoacyl tRNA synthetases. That some mutations are dominant suggests a gain-of-function. Native tRNA synthetases, like TyrRS and TrpRS, catalyze aminoacylation and are also procytokines that are activated by natural fragmentation. In principle, however, gain-of-function phenotypes could arise from mutational activation of synthetase procytokines. From crystal structure analysis we hypothesized that a steric block of a critical ELR motif in full-length TyrRS suppresses the cytokine activity of a natural fragment. To test this hypothesis, we attempted to uncover ELR in the procytokine by mutating a conserved tyrosine (Y341) that tethers ELR. Site-specific proteolytic cleavage and small angle X-ray scattering established subtle opening of the structure by the mutation. Strikingly, four different assays demonstrated mutational activation of cytokine functions. The results prove the possibilities for constitutive gain-of-function mutations in tRNA synthetases.
Aminoacyl-tRNA synthetase; Cytokine function; Tyrosyl-tRNA synthetase; Structure
Pigment epithelium-derived factor (PEDF) was first isolated from the medium conditioned by human fetal retinal pigment epithelial cells and has been detected in a broad range of human fetal and adult tissues. Recent studies have indicated that PEDF activity is inhibitory to angiogenesis.
To study the expression and distribution of pigment epithelium-derived factor (PEDF) in human melanocytes, malignant melanoma cells and tissues.
PEDF was expressed in human melanocytes. The expression of PEDF protein diminished in the following orders healthy skin, pigmented nevus and human malignant melanoma (p < 0.001). Both the expression of PEDF mRNA and protein was much lower or almost absent in the malignant melanoma cell line A375 than that in human melanocytes (p < 0.001).
The expression and distribution of PEDF in human healthy skin, pigmented nevus and malignant melanoma were studied. The expression of PEDF mRNA in human melanocytes and malignant melanoma cell line A375 was measured by reverse transcription polymerase chain reaction (RT-PCR) and PEDF protein was detected by immunohistochemical method and Western blotting analysis.
The lack of PEDF expression may contribute to the pathogenesis of malignant melanoma.
malignant melanoma; pigmented nevus; pigment epithelium derived factor; melanocytes; angiogenesis