Sequencing-free whole-genome spatial transcriptomics at single-molecule resolution.
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Published Erratum↗
Biomedical subjects
Yuan Zhang
Publications and source records attributed to Yuan Zhang.
Haplotype-resolved T2T genome assembly of wild Cannabis provides molecular insights into drought tolerance.
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Letter↗
Therapeutic Targeting of Decr1 Ameliorates Cardiomyopathy by Suppressing Mitochondrial Fatty Acid Oxidation in Diabetic Mice.
BACKGROUND: A significant increase in mitochondrial fatty acid oxidation (FAO) is now increasingly recognized as one of the metabolic alterations in diabetic cardiomyopathy (DCM). However, the molecular mechanisms underlying mitochondrial FAO impairment in DCM remain to be fully elucidated. METHODS: A type 2 diabetes (T2D) mouse model was established by a combination of high-fat diet (HFD) and streptozotocin (STZ) injection. Neonatal rat cardiomyocytes were treated with high glucose (HG) and palmitic acid (HP) to simulate diabetic cardiac injury. Gain- and loss-of-function approaches and RNA sequencing were utilized to investigate the role and mechanism of 2,4-dienoyl-CoA reductase 1 (Decr1) in DCM. RESULTS: By integrating the genomic data available in the Gene Expression Omnibus (GEO) with DCM rodents, we found that the transcriptional level of Decr1 was consistently upregulated in DCM (+255% for diabetic heart, p < 0.0001; +281% for diabetic cells, p < 0.0001). Cardiomyocytes-specific knockdown of Decr1 preserved cardiac function (+41% for EF, p < 0.0001; +24% for FS, p = 0.0052), inhibited cardiac hypertrophy (-34%, p < 0.0001), fibrosis (-69%, p < 0.0001), apoptosis (-56%, p < 0.0001) and oxidative damage (-59%, p < 0.0001) in DCM mice, while cardiomyocytes-specific overexpression of Decr1 aggravated DCM (-28% for EF, p = 0.0347; -17% for FS, p = 0.0014). Deletion of Decr1 prevented high glucose/palmitate (HG/HP)-induced hypertrophy (-22%, p = 0.0006), mitochondrial dysfunction and apoptosis (-74%, p < 0.0001) in cultured cardiomyocytes. Furthermore, RNA sequencing and functional analysis showed that Decr1 interacted with and upregulated pyruvate dehydrogenase kinase 4 (PDK4) in injured cardiomyocytes, and overexpression of PDK4 eliminated the benefits of Decr1 downregulation in DCM (-20% for EF, p = 0.0071; -28% for FS, p = 0.0022). Mechanistically, PDK4 acted as a kinase that induced phosphorylation and mitochondrial translocation of HDAC3. In the mitochondria, HDAC3 mediated the deacetylation of dehydrogenase trifunctional multienzyme complex α subunit (HADHA), contributing to excessive mitochondrial FAO and subsequent cardiac injury. From a screening of 256 natural products, we identified Atranorin and Kurarinone as potential inhibitors of Decr1, both demonstrating protective effects against DCM (Atranorin, +21% for EF, p = 0.0134; +24% for FS, p = 0.0006; Kurarinone, +20% for EF, p = 0.0183; +27% for FS, p = 0.0001). CONCLUSIONS: Our study delineates a molecular mechanism by which Decr1 potentiated higher mitochondrial lipid oxidation and cardiac damage by enhancing HADHA deacetylation through the PDK4/HDAC3 signalling pathway.
Animals↗
Putative rhamnogalacturonan-II glycosyltransferase identified through callus gene editing which bypasses embryo lethality.
Rhamnogalacturonan II (RG-II) is a structurally complex and conserved domain of the pectin present in the primary cell walls of vascular plants. Borate cross-linking of RG-II is required for plants to grow and develop normally. Mutations that alter RG-II structure also affect cross-linking and are lethal or severely impair growth. Thus, few genes involved in RG-II synthesis have been identified. Here, we developed a method to generate viable loss-of-function Arabidopsis (Arabidopsis thaliana) mutants in callus tissue via CRISPR/Cas9-mediated gene editing. We combined this with a candidate gene approach to characterize the male gametophyte defective 2 (MGP2) gene that encodes a putative family GT29 glycosyltransferase. Plants homozygous for this mutation do not survive. We showed that in the callus mutant cell walls, RG-II does not cross-link normally because it lacks 3-deoxy-D-manno-octulosonic acid (Kdo) and thus cannot form the α-L-Rhap-(1→5)-α-D-kdop-(1→sidechain). We suggest that MGP2 encodes an inverting RG-II CMP-β-Kdo transferase (RCKT1). Our discovery provides further insight into the role of sidechains in RG-II dimerization. Our method also provides a viable strategy for further identifying proteins involved in the biosynthesis of RG-II.
Arabidopsis↗
Mechanisms of co-modified liver-targeting liposomes as gene delivery carriers based on cellular uptake and antigens inhibition effect.
In order to deliver antisense oligonucleotides (asODN) into hepatocytes orientedly in the treatment of hepatitis B virus (HBV) infection, the liver-targeting cationic liposomes was developed as a gene carrier, which was co-modified with the ligand of the asialoglycoprotein receptor (ASGPR), beta-sitosterol-beta-d-glucoside (sito-G) and the nonionic surfactant, Brij 35. Flow cytometry (FCM) analysis and enzyme-linked immunosorbent assay (ELISA) showed that the asODN-encapsulating cationic liposomes exhibited high transfection efficiency and strong antigens inhibition effect in primary rat hepatocytes and HepG2.2.15 cells, respectively. With the help of several inhibitors acting on different steps during the targeting lipofection, the cellular uptake mechanisms of the co-modified liver-targeting cationic liposomes were investigated through antigens inhibition effect assay and confocal laser scanning microscopy (CLSM) analysis. The cellular uptake with high transfection efficiency seemed to involve both endocytosis and membrane fusion. The ligand sito-G was confirmed to be able to enhance ASGPR-mediated endocytosis, the nonionic surfactant Brij 35 seemed to be able to facilitate membrane fusion, and the co-modification resulted in the most efficient transfection but no enhanced cytotoxicity. These results suggested that the co-modified liver-targeting cationic liposomes would be a specific and effective carrier to transfer asODN into hepatocytes infected with HBV orientedly.
Androstadienes↗
Puparial case hydrocarbons of Chrysomya megacephala as an indicator of the postmortem interval.
The puparial case is one of the most common 'stages' of necrophagous flies encountered in crime investigations of highly decomposed corpses. If methods for determining the weathering time of these puparial cases are developed, it is possible that the postmortem interval (PMI) could be estimated accordingly. Gas chromatography coupled with mass spectrometry (GC-MS) was used to determine the changes with the weathering time in cuticular hydrocarbons of the puparial cases of Chrysomya megacephala in the laboratory. The results have shown that cuticular hydrocarbons of the puparial cases were a mixture of n-alkanes, methyl-branched alkanes, and dimethyl-branched alkanes. The carbon chain length ranged from C21 to C35, and the hydrocarbon composition showed significant regular changes with the weathering time. For the even numbered n-alkanes with low molecular weight, namely n-C22, n-C24 and n-C26, the abundance increased significantly with the weathering time. For n-C26, in particular, a linear increase in abundance with the weathering time was observed. In addition, for most of the other low molecular weight hydrocarbons (n-C26 or below), the abundance decreased significantly with the weathering time. It is concluded that, cuticular hydrocarbon is a potential indicator of the weathering time in C. megacephala, and possibly in other necrophagous flies, and might further be used to determine the PMI.
Animals↗
Delivery of telomerase reverse transcriptase small interfering RNA in complex with positively charged single-walled carbon nanotubes suppresses tumor growth.
PURPOSE: To determine whether -CONH-(CH(2))(6)-NH(3)(+)Cl(-) functionalized single-walled carbon nanotubes (SWNT) carrying complexed small interfering RNA (siRNA) can enter into tumor cells, wherein they release the siRNA to silence the targeted gene. EXPERIMENTAL DESIGN: -CONH-(CH(2))(6)-NH(3)(+)Cl(-) was used to mediate the conjugation of telomerase reverse transcriptase (TERT) siRNA to SWNTs. The ability of TERT siRNA delivered via SWNT complexes to silence the expression of TERT was assessed by their effects on the proliferation and growth of tumor cells both in vitro and in mouse models. RESULTS: The functionalized SWNTs -CONH-(CH(2))(6)-NH(3)(+)Cl(-) could facilitate the coupling of siRNAs that specifically target murine TERT expression to form the mTERT siRNA:SWNT+ complex. These functionalized SWNTs rapidly entered three cultured murine tumor cell lines, suppressed mTERT expression, and produced growth arrest. Injection of mTERT siRNA:SWNT+ complexes into s.c. Lewis lung tumors reduced tumor growth. Furthermore, human TERT siRNA:SWNT+ complexes also suppressed the growth of human HeLa cells both in vitro and when injected into tumors in nude mice. CONCLUSIONS: -CONH-(CH(2))(6)-NH(3)(+)Cl(-) functionalized SWNTs carry complexed siRNA into tumor cells, wherein they release the siRNA from the nanotube sidewalls to silence the targeted gene. The -CONH-(CH(2))(6)-NH(3)(+)Cl(-) functionalized SWNTs may represent a new class of molecular transporters applicable for siRNA therapeutics.
Animals↗
Oxidized lipid-driven chemokine receptor switch, CCR2 to CX3CR1, mediates adhesion of human macrophages to coronary artery smooth muscle cells through a peroxisome proliferator-activated receptor gamma-dependent pathway.
BACKGROUND: Recent genetic data in mouse and humans suggest that the chemokine receptors CCR2 and CX3CR1 are involved in atherogenesis; however, detailed molecular and cellular mechanisms have not been fully delineated. METHODS AND RESULTS: Here, we show that oxidized linoleic acid metabolites, which are components of oxidized LDL found in large amounts in atherosclerotic plaque, were able to specifically induce differentiation of human monocytes to macrophages with decreased expression of CCR2, confirming a previous report, and increased expression of CX3CR1. These macrophages acquired the ability to adhere to coronary artery smooth muscle cells. The adhesion was mediated directly and predominantly by CX3CR1. Reciprocal effects of these lipids on CCR2 and CX3CR1 expression were mediated by the nuclear receptor peroxisome proliferator-activated receptor (PPAR) gamma, and targeting the PPARgamma gene with sRNAi dramatically reduced macrophage adhesion to coronary artery smooth muscle cells. CONCLUSIONS: These data suggest that in atherogenesis oxidized lipid-driven activation of macrophage PPARgamma in the intima may result in a proadhesive chemokine receptor switch-CCR2 off, CX3CR1 on-causing cessation of CCR2-dependent migration and activation of CX3CR1-dependent retention mechanisms, which together promote macrophage accumulation in vessel wall. Our results may explain at the molecular and cell biology levels the genetic link between CX3CR1 and atherosclerosis. Moreover, they identify macrophage binding to coronary artery smooth muscle cells as the first primary cell setting in which CX3CR1 functions as the major adhesion system.
Atherosclerosis↗
Method for the determination of blood methotrexate by high performance liquid chromatography with online post-column electrochemical oxidation and fluorescence detection.
Methotrexate (MTX) has been widely used at low dose for the treatment of different diseases including rheumatoid arthritis. MTX might be present in plasma in free form, and in blood cells in methotrexate polyglutamate (MTXPG). A rapid and sensitive HPLC method was developed for the determination of plasma MTX level, whole-blood MTX level, and whole-blood total MTX (MTX+MTXPG) level. To determine plasma MTX level or whole-blood MTX level, a 0.2-ml aliquot of plasma or whole blood (after a freeze-thaw cycle to break blood cells) was well mixed with 0.8 ml methanol and centrifuged. To determine whole-blood total MTX level, a 0.1-ml aliquot of whole blood (after a freeze-thaw cycle) was mixed with 80 microl ascorbic acid (114 mM) and incubated at 37 degrees C for 2h to enzymatically convert the MTXPG to MTX. Then 20 microl NaOH solution (0.5M) and 0.8 ml methanol were added and mixed well. After centrifugation, a 0.5-ml aliquot of the supernatant was evaporated to dryness and re-dissolved in 0.2 ml hydrochloric acid (10mM). Methylene chloride (0.2 ml) was added and mixed well. After centrifugation, the top aqueous layer was injected to HPLC for analysis. After the MTX was eluted from the HPLC column, it was electrochemically oxidized and detected by a fluorescence detector. Recoveries of spiked MTX at ppb (ng/ml) level were between 87.9 and 118% with within-day relative standard deviation less than 5.2% and day-to-day relative standard deviation less than 9.8%. The limit of detection (LOD) and limit of quantitation (LOQ) of the described method were 1.2 and 2.6 ng/ml, respectively.
Arthritis, Rheumatoid↗
Identification and characterization of the CD226 gene promoter.
CD226 is one of the main activating receptors on natural killer cells, and it can induce cytotoxicity to target cells through interaction with its ligands CD155 or CD112. CD226 is also involved in T cell differentiation, activation, and cytotoxicity. The expression of CD226 on natural killer cells and T cells can be regulated by cytokines and chemical stimuli; however, the mechanism of the regulation of the CD226 gene is still unknown. In this study, we have identified two promoters in the human CD226 gene named P1 and P2, which are located at -810 to -287 bp and +33 to +213 bp, respectively, and a negative regulation element between P1 and P2. Both P1 and P2 can be regulated by phorbol ester (12-O-tetradecanoylphorbol-13-acetate) and calcium ionophore (A23187). Bioinformatics analysis shows that, within this CD226 gene region, there are putative binding sites for transcription factors AP-1, Sp1, PEA3, and Ets-1. We have found that transcription factor activating protein-1 (AP-1) can up-regulate CD226 promoters P1 and P2 in human hepatocarcinoma cells, a hepatocarcinoma cell line with low expression of endogenous AP-1 and Ets-1. Interestingly, the transcription factor Ets-1 promotes AP-1-induced P2 activity but inhibits AP-1-induced P1 activity for which a 10-bp AP-1/Ets-1 composite site (CCTTCCTTCC) in P1 may be responsible.
Antigens, Differentiation, T-Lymphocyte↗
Sterol intermediates from cholesterol biosynthetic pathway as liver X receptor ligands.
The liver X receptors (LXRs) are ligand-activated transcription factors that regulate the expression of genes controlling lipid metabolism. Oxysterols bind LXRs with high affinity in vitro and are implicated as ligands for the receptor. We showed previously that accumulation of selected dietary sterols, in particular stigmasterol, is associated with activation of LXR in vivo. In the course of the defining of structural features of stigmasterol that confer LXR agonist activity, we determined that the presence of an unsaturated bond in the side chain of the sterol was necessary and sufficient for activity, with the C-24 unsaturated cholesterol precursor sterols desmosterol and zymosterol exerting the largest effects. Desmosterol failed to increase expression of the LXR target gene, ABCA1, in LXRalpha/beta-deficient mouse fibroblasts, but was fully active in cells lacking cholesterol 24-, 25-, and 27-hydroxylase; thus, the effect of desmosterol was LXR-dependent and did not require conversion to a side chain oxysterol. Desmosterol bound to purified LXRalpha and LXRbeta in vitro and supported the recruitment of steroid receptor coactivator 1. Desmosterol also inhibited processing of the sterol response element-binding protein-2 and reduced expression of hydroxymethylglutaryl-CoA reductase. These observations are consistent with specific intermediates in the cholesterol biosynthetic pathway regulating lipid homeostasis through both the LXR and sterol response element-binding protein pathways.
ATP Binding Cassette Transporter 1↗
Genetic characteristics of inbred Wuzhishan miniature pigs, a native Chinese breed.
The Wuzhishan miniature pig originates from Hainan Island, P. R. China, and is considered useful for medical and veterinary research due to its small size; a mature adult weights less than 25 kg. To reveal the genetic characteristics of this breed, the genetic polymorphism in 4 inbred strains was estimated using 30 microsatellite genes, multiplex polymerase chain reaction, and gene scanning. The average number of alleles at the 30 marker loci was 8.300, 4.533, 6.700, and 7.433, respectively. The polymorphism information content was 0.632, 0.507, 0.653, and 0.691, respectively. The average homozygosity in these strains was 0.537, 0.546, 0.549, and 0.520, respectively. These results indicate a relatively high degree of heterozygosity, perhaps becaquse these strains were inbred for 3 generations. Therefore, a long-term inbred breeding program should be conducted for Wuzhishan pigs to establish an experimental model.
Animals↗
CD80 in immune suppression by mouse ovarian carcinoma-associated Gr-1+CD11b+ myeloid cells.
An elevated number of Gr-1+CD11b+ myeloid cells has been described in mice bearing transplantable tumors, and has been associated with immune suppression. We examined the role of such myeloid suppressor cells in mice bearing the spontaneously transformed syngeneic mouse ovarian surface epithelial cell line, 1D8. We observed high levels of CD80 expression by Gr-1+CD11b+ cells from spleen, ascites, and tumor tissue of mice bearing 1D8 ovarian carcinoma, whereas CD40 and CD86 were absent. CD80 expression was not detected on Gr-1+CD11b+ cells from naïve mice. However, the expression of CD80 by Gr-1+CD11b+ cells from naïve mice was promoted by coculture with 1D8 cells. Because irradiated 1D8 cells, but not 1D8-conditioned medium, up-regulate CD80 expression by Gr-1+CD11b+ cells, this phenomenon likely requires direct interaction. Gr-1+CD11b+ cells derived from 1D8 tumor-bearing mice provided significant suppression of antigen-specific immune responses, but Gr-1+CD11b+ cells from naïve mice did not. Both short interfering RNA-mediated knockdown and genetic knockout of CD80 expression by Gr-1+CD11b+ cells of 1D8 tumor-bearing mice alleviated the suppression of antigen-specific immune responses. Suppression via CD80 on Gr-1+CD11b+ myeloid cells was mediated by CD4+CD25+ T regulatory cells and required CD152. CD80 knockout or antibody blockade of either CD80 or CD152 retarded the growth of 1D8 tumor in mice, suggesting that expression of CD80 on Gr-1+CD11b+ myeloid cells triggered by 1D8 ovarian carcinoma suppresses antigen-specific immunity via CD152 signaling and CD4+CD25+ T regulatory cells. Thus, CD80-dependent responses to myeloid suppressor cells may contribute to tumor tolerance and the progression of ovarian carcinoma.
Animals↗
Regulation by nicotine of Gpr51 and Ntrk2 expression in various rat brain regions.
Our previous genetic studies demonstrated that variants of the gamma-Aminobutyric acid B receptor subunit 2 (GPR51) and neurotrophic tyrosine kinase receptor type 2 (NTRK2) genes are significantly associated with nicotine dependence (ND) in smokers. However, whether such genetic associations lead to changes in the expression of the two genes in response to nicotine remains undetermined. In this study, we investigated the regulatory effect of nicotine on the expression of Gpr51 and Ntrk2 in seven rat brain regions during the administration of nicotine in a daily dose of 3.15 mg/kg for 7 days. With quantitative real-time RT-PCR, we found that nicotine increased the mRNA of Gpr51 by 70, 78, and 32% in the amygdala, striatum, and prefrontal cortex (PFC), respectively, but decreased by 54% in the nucleus accumbens (NA). The Gpr51 protein was upregulated by nicotine in the amygdala (26%), striatum (73%), PFC (28%), and medial basal hypothalamus (MBH; 19%) but downregulated in the NA (-72%). Similarly, the mRNA level of Ntrk2 was enhanced by nicotine in the striatum (86%) and PFC (38%), but decreased in the NA (-46%) and ventral tegmental area (VTA; -49%). A significant change in protein expression was also obtained for Ntrk2 in the PFC (24%), MBH (33%), NA (-33%), and VTA (-70%). Interestingly, these two genes showed a closely coordinated expression pattern in response to nicotine in most of the brain regions examined. In summary, our results demonstrate that the expression of Gpr51 and Ntrk2 is significantly regulated by nicotine at both the mRNA and protein levels in various brain regions, which provides further evidence that these two genes are involved in the etiology of ND, as reported in our previous genetic association studies in humans.
Animals↗
Expression of urocortin 2 and its inhibitory effects on intracellular ca2+ via L-type voltage-gated calcium channels in rat pheochromocytoma (PC12) cells.
Urocortin 2, a new member of the corticotrophin-releasing factor (CRF) neuropeptide family, was reported to be widely expressed in the central nervous system and peripheral tissues. Here, we detected urocortin 2 mRNA in PC12 cells using reverse transcription-polymerase chain reaction (RT-PCR). Furthermore, we observed its effects on intracellular Ca(2+) concentration ([Ca(2+)](i)) using confocal microscopy and flow cytometry and on voltage-gated calcium channel (VGCC) currents using whole-cell patch clamp. Our results showed that urocortin 2 mRNA was coexpressed with CRF, and CRF receptor (CRFR) 2beta in undifferentiated PC12 cells, but not CRFR1 or CRFR2alpha. KCl (40 mM) or Bay K8644 (1 microM), an L-type VGCC activator, increased [Ca(2+)](i). Pretreatment of the cells with urocortin 2 significantly diminished the effect of Bay K8644 or KCl. Urocortin 2 showed no influence on [Ca(2+)](i) in tyrode's solution containing EGTA or Ca(2+)-free tyrode's solution. It reversibly inhibited the VGCC currents in a concentration-dependent manner, but had no apparent effects on the cells treated with nifedipine (1 microM), an L-type VGCC blocker. Urocortin 2 up-shifted the current-voltage curves. No frequency-dependence of urocortin 2 effects on I(Ba) was observed. The inhibitory effects of urocortin 2 on VGCC currents or [Ca(2+)](i) were not affected by astressin 2B, an antagonist of CRFR2. As calcium overload play a key role in some neuronal degenerative diseases such as Alzheimer's and Parkinson's diseases, our results suggest that urocortin 2 may be a potentially interesting agent for the treatment of these diseases.
Animals↗
Urocortin II inhibits the apoptosis of mesenteric arterial smooth muscle cells via L-type calcium channels in spontaneously hypertensive rats.
Urocortin (UCN) II, a newly isolated corticotropinreleasing- factor (CRF) related peptide, has been found to have potent cardiovascular protective effects. To investigate the mechanisms of its vascular protective effects, we exposed mesenteric arterial smooth muscle cells (MASMC) from spontaneously hypertensive rats (SHR) to UCN II to observe the change in cell apoptosis using TUNEL assay and measured intracellular calcium concentration ([Ca2+]i) using confocal laser scanning microscope. In addition, effects of UCN II on L-type calcium currents (ICa,L) were also measured using whole-cell patch clamp. Our results showed that UCN II concentration-dependently, but time-independently inhibited cell apoptosis. Astressin 2B, a special CRF 2 receptor antagonist, had no influence on this inhibition. Hypoxia or Bay K8644, the L-type calcium channel activator, induced the apoptosis of MASMC from SHR. Pretreatment of the cells with UCN II diminished the effects of hypoxia or Bay K8644. UCN II was also observed to reduce [Ca2+]i increase induced by KCl or Bay K8644. UCN II concentration-dependently inhibited ICa,L, which was not affected by astressin 2B. It did not affect the activation of ICa,L, but markedly shifted the inactivation curve to the left. In conclusion, UCN II inhibits the apoptosis of MASMC from SHR via inhibiting L-type calcium channels.
3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
Promoter hypermethylation of FANCF plays an important role in the occurrence of ovarian cancer through disrupting Fanconi anemia-BRCA pathway.
OBJECTIVE: Individuals with Fanconi anemia (FA) are predisposed to develop ovarian cancer than those without FA, this is largely contributed to promoter methylation of the FANCF gene and subsequent disruption of the FA-BRCA pathway. In an attempt to understand more molecular genetic bases of ovarian cancer, we examined the expression of the FANCF and the status of the promoter methylation of the FANCF gene in ovarian cancer. METHODS: Seven ovarian cancer cell lines and eighteen ovarian cancer specimens were selected for this study. Both genomic DNA and total RNA were extracted from fresh tissues and cell lines. The DNA was treated with bisulfite and then analyzed with methylation-specific PCR (MSP) to detect FANCF methylation. The expression of FANCF mRNA was detected with Reverse transcription-polymerase chain reaction (RT-PCR). Additionally, the proliferation of cell lines before and after the treatment with demethylating agent 5-Aza-2'-deoxycytidine(5-ADC) was examined with 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide(MTT) assay. RESULTS: The expression of FANCF mRNA decreased in most ovarian cancers as compared with those in normal ovarian tissues. Similarly, the level of FANCF protein decreased in ovarian cancers. The decrease of FANCF was due, in part, to FANCF methylation (five of 12 specimens). The methylation rates were 27.8% (five of 18 specimens) in primary tumors and 14.3% (one of seven cell lines) in established ovarian cancer cell lines respectively. The treatment of ovarian cancer cells with 5-ADC contributed to the following results: the inhibition of DNA promoter methylation, the reactivation of FANCF mRNA expression and protein, and the subsequent reduction in the proliferation of tumor cells both in vitro and in vivo. CONCLUSIONS: The results showed that FANCF methylation regulates the expression of FANCF at both mRNA and protein levels. Methylation-induced inactivation of FANCF plays an important role in the occurrence of ovarian cancers via disrupting the FA-BRCA pathway.
Animals↗
Expression and functional analysis of platelet-derived growth factor in uterine leiomyomata.
OBJECTIVE: To examine the expression of PDGF and its receptors in leiomyoma tissue and to investigate the regulation of PDGF on leiomyoma cell proliferation. MATERIALS AND METHODS: The expression of PDGF and its receptors was examined in 21 pairs of uterine leiomyoma and the adjacent myometrium tissues using an immunostaining method. Paired cultures of leiomyoma and normal myometrium cells were established and treated with PDGF. Total RNA was extracted from leiomyoma and myometrial cells for detection of the expression of proliferating cell nuclear antigen (PCNA) and collagen alpha1 (I, III) by semi-quantitative PCR. RESULT: The expression of PDGF-AA and PDGF-BB in leiomyoma tissue is obviously higher than that in the matched myometrial tissue (P < 0.05). In addition, the expression of PDGF-BB in secretory phase is higher than that in proliferative phase of the menstrual cycle (P = 0.027) in leiomyoma tissue. The expression of PCNA and collagen alpha1 (I) were increased in both leiomyoma and myometrial cells after treatment of PDGF, whereas the expression levels were greater in leiomyoma cells than that in myometrial cells. CONCLUSION: The expression of PDGF and its receptors in leiomyoma and myometrial tissue varied during the menstrual cycle. PDGF may play a role in the pathogenesis of leiomyomas through a mechanism involved in not only the proliferation of leiomyoma cells but also excessive expression of extra cellular molecules.
Becaplermin↗