Humans ; Anemia, Aplastic/genetics* ; Red-Cell Aplasia, Pure ; Chromosome Disorders

Whether Fanconi anemia (FA) heterozygotes are predisposed to bone marrow failure and hematologic neoplasm is a crucial but unsettled issue in cancer prevention and family consulting. We retrospectively analyzed rare possibly significant variations (PSVs) in the five most obligated FA genes, BRCA2, FANCA, FANCC, FANCD2, and FANCG, in 788 patients with aplastic anemia (AA) and hematologic malignancy. Sixty-eight variants were identified in 66 patients (8.38%). FANCA was the most frequently mutated gene (n = 29), followed by BRCA2 (n = 20). Compared with that of the ExAC East Asian dataset, the overall frequency of rare PSVs was higher in our cohort (P = 0.016). BRCA2 PSVs showed higher frequency in acute lymphocytic leukemia (P = 0.038), and FANCA PSVs were significantly enriched in AA and AML subgroups (P = 0.020; P = 0.008). FA-PSV-positive MDS/AML patients had a higher tumor mutation burden, higher rate of cytogenetic abnormalities, less epigenetic regulation, and fewer spliceosome gene mutations than those of FA-PSV-negative MDS/AML patients (P = 0.024, P = 0.029, P = 0.024, and P = 0.013). The overall PSV enrichment in our cohort suggests that heterozygous mutations of FA genes contribute to hematopoietic failure and leukemogenesis.
Anemia, Aplastic/genetics* ; Epigenesis, Genetic ; Fanconi Anemia/genetics* ; Germ Cells ; Hematologic Neoplasms/genetics* ; Humans ; Leukemia, Myeloid, Acute/genetics* ; Retrospective Studies

OBJECTIVE: To explore the effect of miR-335-5p/ADCY3 interaction on the lymphocyte function in the patients with aplastic anemia (AA). METHODS: Blood samples were collected from 22 healthy volunteers (HC) and 50 AA patients including 38 severe AA (SAA) and 12 non-severe AA (NSAA). Peripheral blood mononuclear cells (PBMNC) were isolated. The expression of miR-335-5p and ADCY3 mRNA was detected by using RT-PCR. Negative control miR-335-5p (NC group) and miR-335-5p mimic (mimic group) were transfected to AA-PBMNC by using RNAimax reagent, respectively. The proliferative ability, activation and cytokines of CD4 T and CD8 T cells were measured by flow cytometry. Dual-luciferase reporter assay was used to verify the targeted relationship between miR-335-5p and target gene. RESULTS: The expression of miR-335-5p was significantly downregulated in SAA-PBMNC and NSAA-PBMNC compared with HC-PBMNC (0.08±0.01 vs 0.74±0.10, P＜0.01; 0.17±0.02 vs 0.74±0.10, P＜0.01). Meanwhile, the expression of miR-335-5p in SAA-PBMNC was very statistically significantly lower than that in NSAA-PBMNC (P＜0.01). Compared with NC group, upregulation of miR-335-5p in vitro could significantly inhibited the proliferation of CD4 T and CD8 T cells in AA-PBMNC (P＜0.05 and P＜0.05, respectively). And, upregulating miR-335-5p in AA-PBMNC could significantly inhibited the activation of CD4 and CD8 T cells (P＜0.01 and P＜0.01, respectively). The ratio of CD4TNFα T, CD8IFNγ+T and CD8TNFα T cell by up-regulating the expression of miR-335-5p from AA-PBMNC in vitro was also significantly lower (P＜0.01, P＜0.05 and P＜0.05, respectively). In addition, the expression of ADCY3 was higher in AA-PBMNC than that in HC-PBMNC (1.70±0.15 vs 0.76±0.12, P＜0.01). Furthermore, by means of dual-luciferase reporter assay, the luciferase activity of ADCY3'UTR wildtype could be inhibited by miR-335-5p. CONCLUSIONS The expression of miR-335-5p was significantly downregulated in AA, and that correlates with disease severity. Up-regulating miR-335-5p can correct the hyperimmune status in AA patients by targeting ADCY3. These changes may relates with the strengthen of inhibition for targeted gene ADCY3.
Anemia, Aplastic ; genetics ; CD8-Positive T-Lymphocytes ; Humans ; Leukocytes, Mononuclear ; Lymphocyte Count ; MicroRNAs ; genetics

OBJECTIVE: To explore the relationship between HLA-A, -B, -C, -DRB1, -DQB1 gene polymorphism and aplastic anemia （AA）of 65 cases in Northern China. METHODS: The high resolution genotyping of HLA-A, -B, -C, -DRB1, -DQB1 alleles in 65 AA patients and 772 healthy controls was performed with polymerase chain reaction-sequence specific oligonucleotide (PCR-SSO), the relationship between HLA-A, -B, -C, -DRB1, -DQB1 gene polymorphism and aplastic anemia was analyzed by Pearson Chi-square，Continuity Correction， Two-sided Fisher's Exact Test and Odds Ratio. RESULTS: The HLA-B*1302（10% vs 4.21%）, B*3501（7.69% vs 3.89%）, DRB1* 0701（10% vs 4.73%）, DRB1*0901（19.23% vs 7.58%）, DQB1*0202（9.23% vs 3.76%） gene frequency in AA patients was higher than those in health controls, the difference was statistically significant (P<0.05), the χ were 9.049, 4.336, 6.838, 20.974 and 8.968, OR ratio was 2.528, 2.061, 2.239, 2.904 and 2.605. However, the HLA-A*3303（1.54% vs 6.93%）, DQB1*0302（1.54% vs 6.02%） gene frequency in AA patients was lower than those in healthy controls, the difference was statistically significant (P<0.05), the χ was 5.726 and 4.505, the OR ratio were 0.210 and 0.244. CONCLUSION The polymorphism of HLA-A, -B, -DRB1, -DQB1 alleles is associate with AA in these patient cases, the HLA-B*1302, HLA-B*3501, HLA-DRB1*0701, HLA-DRB1*0901 and HLA-DQB1*0202 may be sensitive genes to AA, while the HLA-A*3303 and HLA-DQB1*0302 may be protective genes on AA.
Alleles ; Anemia, Aplastic ; genetics ; China ; Gene Frequency ; Genetic Predisposition to Disease ; HLA Antigens ; genetics ; Humans ; Polymorphism, Genetic

Adult ; Anemia, Aplastic ; chemically induced ; genetics ; immunology ; Benzene ; poisoning ; Gene Expression ; Genes, T-Cell Receptor beta ; genetics ; Humans ; Male

In order to investigate the levels of stem cell factor (SCF) and its receptor c-kit protein and mRNA in pediatric aplastic anemia (AA) and their relevance to the pathogenesis, immunocytochemical and in situ hybridization were utilized to detect the expression of SCF and its receptor c-kit gene protein and mRNA, respectively in 59 children with AA and 51 normal controls. The relationship between SCF and c-kit and the pathogenesis of AA was analyzed subsequently. The results showed that the positive rate of SCF protein and mRNA expression in children with AA was significantly lower than that in healthy controls (P < 0.05). However, there was no significant difference in the positive rate of c-kit protein and mRNA expression between children with AA and control group (P > 0.05). It was concluded that the expression of SCF is significantly decreased in children with AA, which may be closely associated with the pathogenesis of the AA. c-kit may be unrelated to the development of pediatric AA. Therefore, AA in children may have abnormalities at SCF/c-kit signal transduction levels.
Anemia, Aplastic/etiology ; Anemia, Aplastic/*metabolism ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Receptors, Colony-Stimulating Factor/*biosynthesis ; Receptors, Colony-Stimulating Factor/genetics ; Stem Cell Factor/*biosynthesis ; Stem Cell Factor/genetics

Anemia, Aplastic ; therapy ; Child ; Female ; HLA Antigens ; genetics ; immunology ; Humans ; Mesenchymal Stem Cell Transplantation ; Peripheral Blood Stem Cell Transplantation

This study was aimed to explore the gene expression profile characteristics of T lymphocytes involved in pathogenesis of severe aplastic anemia (SAA) and to predict putative curative drugs for SAA by using biological principle of similarity contrast of gene expression profiles between drugs and diseases. SAA and T lymphocyte were used as key words to search gene expression datasets related to pathogenesis of SAA in public Gene Expression Omnibus (GEO) of NCBI. After significance test, gene expression profiling involved in pathogenesis of SAA were screened and applied to cluster analysis. And then SAA-related gene expression profiles were thrown into pharmacological gene expression datasets of 3000 candidate drugs for similarity analysis and significantly negative correlation was used as a screening criterion for selecting putative curative drugs of SAA. The results showed that only one gene expression dataset was found out, i.e. GSE3807. Computational bioanalysis identified a total of 515 candidate genes of T lymphocyte involved in pathogenesis of SAA, whose expression level exceeded more than 2-fold. Among them, 202 genes were upregulated and 313 genes were downregulated. Cluster analysis showed that those genes belonged to different pathways, including nucleic acid metabolic process, ubiquitin-dependent protein catabolic process, Golgi apparatus protein transport, protein phosphorylation and immunoglobin/major histocompatibility complex. Similarity analysis of gene expression profiles of SAA and drugs showed that hydroxycamptothecin and metformin might have a potential therapeutic efficacy on SAA. It is concluded that by means of novel bioinformatics method, gene expression profiling combined with similarity analysis between disease-related gene expression and pharmacological gene expression profiles may be a novel way of drug screening for SAA.
Anemia, Aplastic ; genetics ; Computational Biology ; Drug Evaluation, Preclinical ; Gene Expression Profiling ; Humans ; Oligonucleotide Array Sequence Analysis ; T-Lymphocytes ; metabolism

OBJECTIVETo analyze the clinical, laboratory characteristics and PIG-A gene mutations in patients of myelodysplastic syndromes (MDS) with PNH clones.

METHODS218 MDS patients diagnosed from August 2013 to August 2015 were analyzed. The PIG-A gene mutations were tested in 13 cases of MDS with PNH clones, 17 cases of AA-PNH and 14 cases of PNH selected contemporaneously by PCR and direct sequencing.

RESULTS13 (5.96%) MDS patients were detected with PNH clones (13/218 cases). 9 patients were treated with cyclosporin A (CsA). Patients showed hematological improvement (HI). There were significant differences between MDS-PNH and PNH patients in terms of granulocyte clone size, red cell clone size and LDH levels [19.2% (1.0%-97.7%) vs 60.2% (3.1%-98.0%), P=0.007; 4.3% (0-67.2%) vs 27.9% (2.5%-83.6%), P=0.026; 246 (89-2014) U/L vs 1137 (195-2239) U/L, P=0.049], while the differences were not statistically significant in patients between MDS-PNH and AA-PNH patients [19.2% (1.0%-97.7%) vs 23.2% (1.5%-96.0%), P=0.843; 4.3% (0-67.2%) vs 14.4% (1.1%-62.8%), P=0.079; 246 (89-2014) U/L vs 406 (192-1148) U/L, P=0.107]. PIG-A gene mutations were detected in 7 MDS-PNH patients, of them, six were missense mutations, one were frameshift mutation and four cases with the same mutation of c.356G>A (R119Q). The PIG-A gene mutations were also detected in 9/11 AA-PNH patients and 11/14 PNH patients, both of them had the mutation of c.356G>A (R119Q). The PIG-A gene mutations of MDS-PNH, AA-PNH, PNH patients were all small mutations, the majority of those (59%) were missense mutation and mainly located in exon 2.

CONCLUSIONMDS patients with PNH clones had better response to CsA, smaller PNH clone size. The PIG-A gene mutations of MDS-PNH patients mainly located in exon 2, which could be a mutational hotspot of these patients.

Anemia, Aplastic ; genetics ; Clone Cells ; Erythrocytes ; cytology ; Exons ; Granulocytes ; cytology ; Hemoglobinuria, Paroxysmal ; genetics ; Humans ; Membrane Proteins ; genetics ; Mutation ; Myelodysplastic Syndromes ; genetics ; Polymerase Chain Reaction

Anemia, Aplastic ; diagnosis ; genetics ; Diagnosis, Differential ; HLA Antigens ; genetics ; HLA-DQ Antigens ; genetics ; HLA-DQ beta-Chains ; Humans ; Medicine, Chinese Traditional ; Polymorphism, Genetic ; Somatotypes ; genetics