Usher syndrome:Genetic diagnosis and current therapeutic approaches

Usher Syndrome(USH)is the most common deaf-blind syndrome,affecting approximately 1 in 6000 people in the deaf population.This genetic condition is characterized by a combination of hearing loss(HL),retinitis pigmentosa,and,in some cases,vestibular areflexia.Among the subtypes of USH,USH type 1 is considered the most severe form,presenting profound bilateral congenital deafness,vestibular areflexia,and early onset RP.USH type 2 is the most common form,exhibiting congenital moderate to severe HL for low frequencies and severe to profound HL for high frequencies.Conversely,type 3 is the rarest,initially manifesting mild symptoms during childhood that become more prominent in the first decades of life.The dual impact of USH on both visual and auditory senses significantly impairs patients'quality of life,restricting their daily activities and interactions with society.To date,9 genes have been confirmed so far for USH:MYO7A,USH1C,CDH23,PCDH15,USH1G,USH2A,ADGRV1,WHRN and CLRN1.These genes are inherited in an autosomal recessive manner and encode proteins expressed in the inner ear and retina,leading to functional loss.Although non-genetic methods can assist in patient triage and disease extension evaluation,genetic and molecular tests play a pivotal role in providing genetic counseling,enabling appropriate gene therapy,and facilitating timely cochlear implantation(CI).The CRISPR/Cas9 system and viral-based gene replacement therapy have recently emerged as highly promising techniques for treating USH.Regarding drug therapy,PTC-124 and Nb54 have been identified as promising drug interventions for genetic HL in USH.Simultaneously,CI has proven to be critical in the restoration of hearing.This review aims to summarize the genetic and molecular diagnosis of USH and highlight the importance of early diagnosis in Cuzzuol BR et al.Diagnosis and current treatments of USH WJO https://www.wjgnet.com 2 January 19,2024 Volume 11 Issue 1 guiding appropriate treatment strategies and improving patient prognosis.

EARLY DIAGNOSIS OF MYELODYSPLASTIC SYNDROMES USING CLONAL ANALYSES

Objective: To study the value of clonal analysis to the early diagnosis of myelodysplastic syndrome (MDS). Methods: Four types of clonal analyses were performed on the bone marrow samples from 50 patients suspected of MDS: (1) Conventional Cytogenetics (CC) for clonal chromosomal abnormalities; (2) BrdU-Sister Chromatid Differentiation (BrdU-SCD) for cell cycle kinetics; (3) Fluorescence in Situ Hybridization (FISH) for trisomy 8; (4) Polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) for N-ras mutation. Results: The diagnosis of forty-three patients was compatible with the FAB criteria for MDS. The other seven cases didn’t meet the FAB criteria, with only one lineage of dyspoiesis or with no obvious dysplastic changes. Among these seven cases, two were morphologically diagnosed with suspicious refractory anemia, one with sideroblastic anemia, one with leukemoid reaction, one with hypercellular anemia and two with chronic aplastic anemia. Clonal analyses of the 7 patients showed that six cases had clonal karyotype abnormalities, four had prolonged cell cycle patterns, four had trisomy 8 of different proportions and one had mutation of the exon 1 of N-RAS. Thus, they were revaluated as MDS patients. Conclusion: The untypical MDS patients with one lineage dyspoiesis or without obvious dysplastic changes can be diagnosed early by combining multiple clonal analysis techniques such as CC, SCD, FISH and PCR-SSCR.

Decreased CD10-positive granulocytes for the differential diagnosis of myelodysplastic syndrome

Myelodysplastic syndromes(MDS)are highly heterogeneous myeloid neoplasms,and a large number of patients are difficult to diagnose and classify by blood and bone marrow examination.As a surface marker of granulocyte,studies have shown CD10 can be used to define the degree of granulocyte maturation in MDS patients.However,whether it can be used for differential diagnosis of MDS and other hematological diseases remains inconclusive.To explore the value of CD10 for differential diagnosis of MDS,60 newly diagnosed MDS,20 aplastic anemia(AA)patients,and 35 iron-deficient anemia(IDA)patients were selected for this study.Bone marrow(BM)specimens were processed for surface marker analysis and labeled with pre-conjugated monoclonal antibodies.Stained cells were detected by flow cytometry.Our results indicated that CD10-positive granulocytes were significantly decreased in BM of MDS patients than AA and IDA patients,and the level of CD10-positive mature granulocytes was not associated with the clinical stages of malignancy.Receiver operating characteristic(ROC)areas under the curve(AUC)of CD10-positive granulocytes was 0.86 and 0.85,respectively,in MDS patients than the IDA group and AA group with good specificity and sensitivity.Further,CD10-positive granulocytes were increased after effective treatment.In conclusion,we found the decrease in CD10-positive granulocytes has a differential diagnostic value of MDS.

Recent advances in pathophysiology,diagnosis and management of hepatorenal syndrome:A review

Hepatorenal syndrome with acute kidney injury(HRS-AKI)is a form of rapidly progressive kidney dysfunction in patients with decompensated cirrhosis and/or acute severe liver injury such as acute liver failure.Current data suggest that HRS-AKI occurs secondary to circulatory dysfunction characterized by marked splanchnic vasodilation,leading to reduction of effective arterial blood volume and glomerular filtration rate.Thus,volume expansion and splanchnic vasoconstriction constitute the mainstay of medical therapy.However,a significant proportion of patients do not respond to medical management.These patients often require renal replacement therapy and may be eligible for liver or combined liver-kidney transplantation.Although there have been advances in the management of patients with HRS-AKI including novel biomarkers and medications,better-calibrated studies,more widely available biomarkers,and improved prognostic models are sorely needed to further improve diagnosis and treatment of HRS-AKI.

miR-103-3p regulates the differentiation of bone marrow mesenchymal stem cells in myelodysplastic syndrome

The pathogenesis of myelodysplastic syndrome(MDS)may be related to the abnormal expression of microRNAs(miRNAs),which could influence the differentiation capacity of mesenchymal stem cells(MSCs)towards adipogenic and osteogenic lineages.In this study,exosomes from bone marrow plasma were successfully extracted and identified.Assessment of miR-103-3p expression in exosomes isolated from BM in 34 MDS patients and 10 controls revealed its 0.52-fold downregulation in patients with MDS compared with controls(NOR)and was downregulated 0.55-fold in MDS-MSCs compared with NOR-MSCs.Transfection of MDS-MSCs with the miR-103-3p mimic improved osteogenic differentiation and decreased adipogenic differentiation in vitro,while inhibition of miR-103-3p showed the opposite results in NOR-MSCs.Thus,the expression of miR-103-3p decreases in MDS BM plasma and MDS-MSCs,significantly impacting MDS-MSCs differentiation.The miR-103-3p mimics may boost MDS-MSCs osteogenic differentiation while weakening lipid differentiation,thereby providing possible target for the treatment of MDS pathogenesis.

Severe inflammatory disorder in trisomy 8 without myelodysplastic syndrome and response to methylprednisolone:A case report

BACKGROUND Patients with trisomy 8 consistently present with myeloid neoplasms and/or auto-inflammatory syndrome.A possible link between myelodysplastic syndromes(MDS)with trisomy 8(+8-MDS)and inflammatory disorders is well recognized,several cases having been reported.However,inflammatory disorders in patients without MDS have been largely overlooked.Generally,Behçet's disease is the most common type in+8-MDS.However,inflammatory disorders with pulmonary involvement are less frequent,and no effective treatment has been established.CASE SUMMARY A 27-year-old man with recurrent fever,fatigue for>2 mo,and unconsciousness for 1 day was admitted to our emergency department with a provisional diagnosis of severe pneumonia.Vancomycin and imipenem were administered and sputum collected for metagenomic next-generation sequencing.Epstein–Barr virus and Mycobacterium kansasii were detected.Additionally,chromosomal analysis showed duplications on chromosome 8.Two days later,repeat metagenomic next-generation sequencing was performed with blood culture.Cordyceps portugal,M.kansasii,and Candida portugal were detected,and duplications on chromosome 8 confirmed.Suspecting hematological disease,we aspirated a bone marrow sample from the iliac spine,examination of which showed evidence of infection.We added fluconazole as further antibiotic therapy.Seven days later,the patient’s condition had not improved,prompting addition of methylprednisolone as an anti-inflammatory agent.Fortunately,this treatment was effective and the patient eventually recovered.CONCLUSION Severe inflammatory disorders with pulmonary involvement can occur in patients with trisomy 8.Methylprednisolone may be an effective treatment.

Flared inflammatory episode transforms advanced myelodysplastic syndrome into aplastic pancytopenia:A case report and literature review

BACKGROUND Myelodysplastic syndrome(MDS)is a hematological neoplasm,and an increase in myeloblasts is representative of leukemic hematopoiesis in advanced MDS.Lowrisk MDS usually exhibits deranged autoimmunity resembling that of aplastic anemia(AA),whereas advanced MDS is characterized by a phenotype of immune exhaustion.MDS can be normo/hyperplastic or hypoplastic.Generally,bone marrow cellularity and myeloblasts increase with disease progression.Transformation from advanced MDS to AA-like syndrome with leukemic cell regression has not previously been reported.CASE SUMMARY A middle-aged Chinese woman had a 4-year history of leukocytopenia.Six months prior to admission,the patient developed gradually worsening fatigue and performance status.The leukocytopenia further progressed.She was diagnosed with MDS with excess blasts-2 based on increased bone marrow cellularity and an increased percentage of myeloblasts on marrow and blood smears,an increased percentage of cluster of differentiation(CD)34+CD33+progenitors in immunotyping analysis,a normal karyotype in cytogenetic analysis,and the identification of somatic mutations in CBL,KMT2D and NF1 in molecular analysis.Initially,neutropenia was the predominant hematological abnormality,with mild anemia and thrombocytosis,and the degree of fatigue was far more severe than the degree of anemia.In the following months,the patient experienced several febrile episodes.Intravenous antibiotic treatments were able to control the febrile episodes,but the elevated inflammatory indices persisted.The hematological parameters dramatically fluctuated with the waxing and waning of the inflammatory episodes.With recurrent flares of the inflammatory condition,agranulocytosis and severe anemia developed,with mild thrombocytopenia.During the patient’s hospitalization,computed tomography(CT)scans revealed the presence of extensive inflammatory lesions involving the lungs,mediastinum,pleura,gastrointestinal tract,peritoneum and urinary tract,with imaging features suggestive of the reactivation of disseminated tuberculosis.Reevaluation of the bone marrow smears revealed that the cellularity became hypoplastic,and the leukemic cells regressed,suggesting that both normal and leukemic hematopoiesis had been heavily suppressed.Immunological analysis of the bone marrow samples revealed a decreased percentage of CD34+cells and an immunological signature resembling that of severe AA(SAA),confirming the regression of the leukemic cells by autoimmune-mediated attacks.The patient demonstrated resistance to multiple drugs,including antituberculotics,recombinant human granulocyte colony-stimulating factor,broad-spectrum antibiotics,voriconazole,ganciclovir,immune suppressants,eltrombopag and intravenous immunoglobulin,which further worsened the hematological injury and patient’s performance status.The patient eventually died of overwhelming infection and multidrug resistance.CONCLUSION Advanced MDS can transform to aplastic cytopenia with leukemic cell regression and an immunological signature of SAA during inflammatory flare-ups.

Tuberculosis-induced aplastic crisis and atypical lymphocyte expansion in advanced myelodysplastic syndrome:A case report and review of literature

BACKGROUND Myelodysplastic syndrome(MDS)is caused by malignant proliferation and ineffective hematopoiesis.Oncogenic somatic mutations and increased apoptosis,necroptosis and pyroptosis lead to the accumulation of earlier hematopoietic progenitors and impaired productivity of mature blood cells.An increased percentage of myeloblasts and the presence of unfavorable somatic mutations are signs of leukemic hematopoiesis and indicators of entrance into an advanced stage.Bone marrow cellularity and myeloblasts usually increase with disease progression.However,aplastic crisis occasionally occurs in advanced MDS.CASE SUMMARY A 72-year-old male patient was definitively diagnosed with MDS with excess blasts-1(MDS-EB-1)based on an increase in the percentages of myeloblasts and cluster of differentiation(CD)34+hematopoietic progenitors and the identification of myeloid neoplasm-associated somatic mutations in bone marrow samples.The patient was treated with hypomethylation therapy and was able to maintain a steady disease state for 2 years.In the treatment process,the advanced MDS patient experienced an episode of progressive pancytopenia and bone marrow aplasia.During the aplastic crisis,the bone marrow was infiltrated with sparsely distributed atypical lymphocytes.Surprisingly,the leukemic cells disappeared.Immunological analysis revealed that the atypical lymphocytes expressed a high frequency of CD3,CD5,CD8,CD16,CD56 and CD57,suggesting the activation of autoimmune cytotoxic T-lymphocytes and natural killer(NK)/NKT cells that suppressed both normal and leukemic hematopoiesis.Elevated serum levels of inflammatory cytokines,including interleukin(IL)-6,interferon-gamma(IFN-γ)and tumor necrosis factor-alpha(TNF-α),confirmed the deranged type I immune responses.This morphological and immunological signature led to the diagnosis of severe aplastic anemia secondary to large granule lymphocyte leukemia.Disseminated tuberculosis was suspected upon radiological examinations in the search for an inflammatory niche.Antituberculosis treatment led to reversion of the aplastic crisis,disappearance of the atypical lymphocytes,increased marrow cellularity and 2 mo of hematological remission,providing strong evidence that disseminated tuberculosis was responsible for the development of the aplastic crisis,the regression of leukemic cells and the activation of CD56+atypical lymphocytes.Reinstitution of hypomethylation therapy in the following 19 mo allowed the patient to maintain a steady disease state.However,the patient transformed the disease phenotype into acute myeloid leukemia and eventually died of disease progression and an overwhelming infectious episode.CONCLUSION Disseminated tuberculosis can induce CD56+lymphocyte infiltration in the bone marrow and in turn suppress both normal and leukemic hematopoiesis,resulting in the development of aplastic crisis and leukemic cell regression.

Clinical management of myelodysplastic syndrome/myeloproliferative neoplasm overlap syndromes

The myelodysplastic/myeloproliferative neoplasms(MDS/MPNs) are a unique group of hematologic malignancies characterized by concomitant myelodysplastic and myeloproliferative features. According to the 2008 WHO classification, the category includes atypical chronic myeloid leukemia(a CML), chronic myelomonocytic leukemia(CMML), juvenile myelomonocytic leukemia(JMML), MDS/MPN-unclassifiable(MDS/MPN-U), and the provisional entity refractory anemia with ring sideroblasts and thrombocytosis(RARS-T). Although diagnosis currently remains based on clinicopathologic features, the incorporation of nextgeneration platforms has allowed for the recent molecular characterization of these diseases which has revealed unique and complex mutational profiles that support their distinct biology and is anticipated to soon play an integral role in diagnosis,prognostication, and treatment. Future goals of research should include the development of disease-modifying therapies, and further genetic understanding of the category will likely form the foundation of these efforts.

Identification of SNP-containing regulatory motifs in the myelodysplastic syndromes model using SNP arrays and gene expression arrays

Myelodysplastic syndromes have increased in frequency and incidence in the American population, but patient prognosis has not significantly improved over the last decade. Such improvements could be realized if biomarkers for accurate diagnosis and prognostic stratification were successfully identified. In this study, we propose a method that associates two state-of-the-art array technologies-single nucleotide polymorphism (SNP) array and gene expression array-with gene motifs considered transcription factor -binding sites (TFBS). We are particularly interested in SNP-containing motifs introduced by genetic variation and mutation as TFBS. The potential regulation of SNP-containing motifs affects only when certain mutations occur. These motifs can be identified from a group of co-expressed genes with copy number variation. Then, we used a sliding window to identify motif candidates near SNPs on gene sequences. The candidates were filtered by coarse thresholding and fine statistical testing. Using the regression-based LARS-EN algorithm and a level-wise sequence combination procedure, we identified 28 SNP-containing motifs as candidate TFBS. We confirmed 21 of the 28 motifs with ChIP-chip fragments in the TRANSFAC database. Another six motifs were validated by TRANSFAC via searching binding fragments on coregulated genes. The identified motifs and their location genes can be considered potential biomarkers for myelodysplastic syndromes. Thus, our proposed method, a novel strategy for associating two data categories, is capable of integrating information from different sources to identify reliable candidate regulatory SNP-containing motifs introduced by genetic variation and mutation.

TET2 Expression in Bone Marrow Mononuclear Cells of Patients with Myelodysplastic Syndromes and Its Clinical Significances

Objective To investigate the expression of TET2 mRNA and protein in the bone marrow mononuclear cells(BMMNC) of patients with myelodysplastic syndrome(MDS) and its clinical significance. Methods The expression of TET2 mRNA and protein in bone marrow mononuclear cells(BMMNC) of 32 patients with MDS and 20 healthy donors was examined by qPCR and Western blot. Results The expression of TET2 mRNA in BMMNC was down-regulated in MDS patients compared with the donor group [(0.41±0.28)%vs.(1.07±0.56)%](P<0.001).Compared with lower expression group(TET2<0.4)[(6.53±6.17)%],patients with higher expression of TET2(≥0.4) presented significantly lower proportion of bone marrow blasts[(1.21±1.56)%](P<0.05).The expression of TET2 mRNA in BMMNC of MDS patients was inversely correlated with malignant clone burden(r=-0.398,P<0.05) and IPSS(r=-0.412, P<0.05).The expression of TET2 protein was down-regulated in MDS patients compared with that in the donor group. Conclusions The mRNA and protein expression of TET2 in BMMNC of MDS patients is decreased,which might be useful as an important parameter for the evaluation of MDS clone burden.

BURDEN OF ABNORMAL HEMATOPOIETIC CLONE IN PATIENTS WITH MYELODYSPLASTIC SYNDROMES

Objective To investigate the role of the burden of abnormal hematopoietic clone in the development of myelodysplastic syndromes (MDS). Methods The ratio of the bone marrow cells with abnormal chromosomes to the total counted bone marrow cells was regarded as the index of MDS clone burden. The disease severity related parameters including white blood cell count, hemoglobin, platelet count, lactate dehydrogenase level, bone marrow blast, myeloid differentiation index, micromegakaryocyte, transfusion, interleukin-2, tumor necrosis factor (TNF), CD4^+ and CD8^+ T cells of MDS patients were assayed, and the correlations between those parameters and MDS clone burden were also analyzed. Results The clone burden of MDS patients was 67.4%±36.2%. MDS clone burden positively correlated with bone marrow blasts (r = 0.483, P<0.05), negatively with hemoglobin level (r=-0.445, P<0.05). The number of blasts, hemoglobin, and erythrocytes in high clone burden (>50%) and low clone burden (≤50%) groups were 7.78%±5.51% and 3.45%±3.34%, 56.06±14.28 g/L and 76.40±24.44 g/L, (1.82±0.48)×10~ 12 /L and (2.32±0.66)×10~ 12 /L, respectively (all P<0.05). CD4^+ T lymphocytes of MDS patients and normal controls were (0.274±0.719)×10~ 9 /L and (0.455±0.206)×10~ 9 /L, respectively (P<0.05). CD8^+ T lymphocytes of MDS patients and normal controls were (0.240±0.150)×10~ 9 /L and (0.305±0.145)×10~ 9 /L, respectively. The serum level of interleukin-2 of MDS patients (6.29±3.58 ng/mL) was significantly higher than normal control (3.11±1.40 ng/mL, P<0.05). The serum level of TNF of MDS patients and normal control group were 2.42±1.79 ng/mL and 1.68±0.69 ng/mL, respectively. The ratio of CD4 to CD8 was higher in high clone burden MDS patients (1.90±0.52) than that in low clone burden patients (0.97±0.44, P<0.05). Conclusion The quantitive clonal karyotype abnormalities and deficient T cell immunity are important parameters for evaluating MDS severity and predicting its progression.

Expression of DLK1 Gene in the Bone Marrow Cells of Patients with Myelodysplastic Syndromes and Its Clinical Significance

Objective This study aims to investigate the expression of delta-like 1(DLKl) gene in the bone marrow cells of patients with myelodysplastic syndromes(MDS) and to explore its molecular characteristics for the early diagnosis of MDS. Methods The expression of DLK1 mRNA in the bone marrow cells of cases with MDS,acute myeloid leukemia(AML),and normal control groups were measured by real-time polymerase chain reaction and were analyzed for clinical significance. Results Significantly higher expression of DLK1 mRNA was observed in the bone marrow cells of MDS patients(0.7342±0.3652) compared with the normal control group(0.4801±0.1759)(P<0.05).The expression of DLK1 mRNA had a positive correlation with the proportion of bone marrow blasts(r=0.467,P<0.05).Moreover,DLKl mRNA expression was significantly increased as MDS progressed (P<0.05).Patients with abnormal karyotypes exhibited significantly higher expression of DLKl mRNA(0.9007±0.4334) than those with normal karyotypes(0.6411±0.2630)(P<0.05).Subsequently,patients with highly expressed DLKl(>0.8) presented significantly higher malignant clone burden(0.4134±0.3999) than those with lower DLKl expression(<0.8),(0.1517±0.3109),(P<0.05). Conclusions The DLK1 gene was highly expressed in MDS patients,and was increased as MDS progressed.The expression of DLK1 mRNA was positively correlated with the proportion of the bone marrow blasts.A high expression of DLKl gene suggested a higher malignant clone burden of MDS.

Insights into myelodysplastic syndromes from current preclinical models

In recent years, there has been significant progress made in our understanding of the molecular genetics of myelodysplastic syndromes(MDS). Using massively parallel sequencing techniques, recurring mutations are identified in up to 80% of MDS cases, including many with a normal karyotype. The differential role of some of these mutations in the initiation and progression of MDS is starting to be elucidated. Engineering candidate genes in mice to model MDS has contributed to recent insights into this complex disease. In this review, we examine currently available mouse models, with detailed discussion of selected models. Finally, we highlight some advances made in our understanding of MDS biology, and conclude with discussions of questions that remain unanswered.

Emerging immunological concepts in the pathogenesis of myelodysplastic syndromes

The involvement of T-lymphocytes in the pathogenesis of myelodysplastic syndromes(MDS) is now well documented by relevant clinical and experimental findings.This brief review will focus on the T-cell repertoire pattern typical of MDS patients as well as on the potential role exerted by specific T-cell subsets in this context.Future investigations should further explore the specific role played by different T-cell subsets in the bone marrow milieu typical of MDS, further clarifying which of the described changes represent either an epiphenomenon or rather a real causative factor in the pathogenesis of these disorders.

Remarkably different results between two studies from North America on genomic mutations and sensitivity to DNA demethylating agents for myelodysplastic syndromes

Sekeres et al. （1） conducted a multicenter randomized, controlled trial to compare whether azacitidine-based combinations with lenalidomide or vorinostat produce superior overall response rates to azacitidine in the treatment of myelodysplastic syndromes （MDS）. In that trial, 224 patients with higher-risk MDS and 53 with chronic myelomonocytic leukemia （CMML） were enrolled and randomly assigned to the ＂azacitidine＂ group, ＂azacitidine plus lenalidomide＂ group or ＂azacitidine plus vorinostat＂ group. The researchers found that patients with MDS treated with azacitidine-based combinations had similar response rate to azacitidine monotherapy. Using genomic mutation analysis, they found that the overall response rate to azacitidine-based treatment was higher for patients with mutations in DNMT3A and lower for those with mutations in SRSF2. Whereas in another study, Welch et al. enrolled 26 patients with MDS and 90 with acute myeloid leukemia （AML） who were treated with decitabine, and they found that patients with TP53 mutations had a higher response rate, but not those with DNMT3A mutations （2）. We propose that this big discrepancy in the conclusions between the two studies might have been caused by the presence of many co-interacting factors, e.g. study aims, DNA demethylating agents, treatment protocols, and patient sources.

A case of myelodysplastic syndromes with initial symptom of erythra

Myelodysplastic syndrome(MDS)is a malignant clonal disease of human hematopoietic stem cells.In this paper,a case of MDS with fever and rash as the first symptom was reported in our hospital,and the related literature was reviewed and summarized to provide ideas for the diagnosis of MDS.Case summary:The patient,male,62 years old,with systemic rash and fever as the initial symptoms,early multiple bone marrow examination showed no typical abnormalities.With the progress of the disease,bone marrow cytology,flow cytometry,molecular karyotype,chromosomal karyotype,skin biopsy and pathological diagnosis were performed,and the diagnosis was MDS.

Variants of the Mitochondrial Displacement Loop in Patients with Myelodysplastic Syndromes

OBJECTIVE Some mtDNA mutations have been detected in patients with myelodysplastic syndromes(MDSs).As the non-coding region of mitochondria,the displacement loop(D-loop) region of mtDNA contains important elements for mtDNA replication and transcription.Variants of the D-loop region were found to be related to the cause of many diseases.The aim of our study was to investigate mutations and single nucleotide polymorphisms in the D-loop region of MDS patients.METHODS The mutations and SNPs in the hypervariable regions of the D-loop were detected by direct sequencing in MDS patients and normal controls.RESULTS Sixty-four SNPs were found in the D-loop region in MDS cases and control group.Among the SNPs,the 16,189 variant(T > C transition) was found to have an increased frequency in the MDS group(P = 0.044).However,no mutations were detected in neither group.CONCLUSION Our data provide evidence for a highly polymorphic D-loop region in patients with MDS,but do not support the presence of mutations in the mitochondrial D-loop region in MDS cases.The mtDNA T16,189C variant,which may be a functional variant,is associated with increased susceptibility to a MDS.

Exploration of the molecular mechanism of Indigo Naturalis in the treatment of myelodysplastic syndromes through integrated pharmacological study

Background:Oral administration of indigo naturalis(IN)is used as a complementary and alternative medicine(CAM)regimen for the treatment of myelodysplastic syndromes(MDS).However,its mechanism of action has not been fully elucidated and needs to be further explored.Methods:By searching the traditional Chinese medicine system and analyzing platforms(TCMSP),bioinformatics analysis tool for the molecular mechanism of traditional Chinese medicine(BATMAN-TCM),and Swiss Target Prediction network database,the main active components and potential targets of IN were obtained.Based on this,a component-target network was established by Cytoscape 3.6.1 software.Differentially expressed genes(DGEs)in MDS were obtained from three GEO(Gene Expression Omnibus)gene chips.Then,the protein-protein interaction(PPI)network of DGEs was constructed and analyzed by STRING database and Cytoscape 3.6.1 software.In addition,Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)biological enrichment analysis were carried out using REVIGO and KEGG Orthology Based Annotation System(KOBAS)on DGEs,respectively.Identification of IN-MDS compound targets was performed by matching potential targets of active components with disease-related targets.The results of KEGG pathway enrichment analysis were combined with compound targets to screen key targets.In the end,molecular docking was performed by SYBYL-X2.1 to verify the key targets.Results:Nine active components of IN and 439 potential targets of IN were identified by analyzing TCMSP,BATMAN-TCM,and Swiss Target Prediction network databases.Three MDS disease-related gene microarray chips were obtained from the GEO databases:GSE4619,GSE19429,and GSE58831.Through this analysis,87 DEGs were finally obtained using the Venn diagram.A PPI network of DEGs was then constructed,in which 18 genes were upregulated and 69 genes were downregulated.After the GO enrichment results were de-redundant,the representative GO terms were obtained by using REVIGO semantic similarity measuremen.The KEGG biological pathway analysis using the KOBAS indicated that the Hippo signaling pathway is important in MDS.The Hippo signaling pathway involves four genes:AREG,LEF1,SMAD7,and TCF4.By matching and mapping DEGs with potential targets,six IN-MDS compound targets were obtained:PDE4B,PLAUR,ELANE,NR3C1,AREG,and LEF1.We found that AREG and LEF1 are consistent with the genes involved in the Hippo signaling pathway.Through molecular docking simulation,we found that the indican binds best to AREG and LEF1.Conclusion:Based on the integrated pharmacology model,the material basis of the efficacy and biological molecular mechanism of IN in the treatment of MDS was systematically studied,which provided a novel indication of the CAM regimen for the improvement of MDS management.