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1.
Mol Cell ; 84(14): 2648-2664.e10, 2024 Jul 25.
Article in English | MEDLINE | ID: mdl-38955181

ABSTRACT

The essential Mediator (MED) coactivator complex plays a well-understood role in regulation of basal transcription in all eukaryotes, but the mechanism underlying its role in activator-dependent transcription remains unknown. We investigated modulation of metazoan MED interaction with RNA polymerase II (RNA Pol II) by antagonistic effects of the MED26 subunit and the CDK8 kinase module (CKM). Biochemical analysis of CKM-MED showed that the CKM blocks binding of the RNA Pol II carboxy-terminal domain (CTD), preventing RNA Pol II interaction. This restriction is eliminated by nuclear receptor (NR) binding to CKM-MED, which enables CTD binding in a MED26-dependent manner. Cryoelectron microscopy (cryo-EM) and crosslinking-mass spectrometry (XL-MS) revealed that the structural basis for modulation of CTD interaction with MED relates to a large intrinsically disordered region (IDR) in CKM subunit MED13 that blocks MED26 and CTD interaction with MED but is repositioned upon NR binding. Hence, NRs can control transcription initiation by priming CKM-MED for MED26-dependent RNA Pol II interaction.


Subject(s)
Cryoelectron Microscopy , Cyclin-Dependent Kinase 8 , Mediator Complex , Protein Binding , RNA Polymerase II , RNA Polymerase II/metabolism , RNA Polymerase II/genetics , Mediator Complex/metabolism , Mediator Complex/genetics , Mediator Complex/chemistry , Humans , Cyclin-Dependent Kinase 8/metabolism , Cyclin-Dependent Kinase 8/genetics , Animals , Intrinsically Disordered Proteins/metabolism , Intrinsically Disordered Proteins/genetics , Intrinsically Disordered Proteins/chemistry , Binding Sites , Receptors, Cytoplasmic and Nuclear/metabolism , Receptors, Cytoplasmic and Nuclear/genetics , HEK293 Cells , Protein Interaction Domains and Motifs
2.
Nucleic Acids Res ; 2024 Aug 24.
Article in English | MEDLINE | ID: mdl-39180400

ABSTRACT

Processing of RNA is a key regulatory mechanism for all living systems. Escherichia coli protein YicC belongs to the well-conserved YicC family and has been identified as a novel ribonuclease. Here, we report a 2.8-Å-resolution crystal structure of the E. coli YicC apo protein and a 3.2-Å-cryo-EM structure of YicC bound to an RNA substrate. The apo YicC forms a dimer of trimers with a large open channel. In the RNA-bound form, the top trimer of YicC rotates nearly 70° and closes the RNA substrate inside the cavity to form a clamshell-pearl conformation that resembles no other known RNases. The structural information combined with mass spectrometry and biochemical data identified cleavage on the upstream side of an RNA hairpin. Mutagenesis studies demonstrated that the previously uncharacterized domain, DUF1732, is critical in both RNA binding and catalysis. These studies shed light on the mechanism of the previously unexplored YicC RNase family.

3.
Nucleic Acids Res ; 51(10): 4814-4830, 2023 06 09.
Article in English | MEDLINE | ID: mdl-36928138

ABSTRACT

The Paf1 complex (Paf1C) is a conserved transcription elongation factor that regulates transcription elongation efficiency, facilitates co-transcriptional histone modifications, and impacts molecular processes linked to RNA synthesis, such as polyA site selection. Coupling of the activities of Paf1C to transcription elongation requires its association with RNA polymerase II (Pol II). Mutational studies in yeast identified Paf1C subunits Cdc73 and Rtf1 as important mediators of Paf1C association with Pol II on active genes. While the interaction between Rtf1 and the general elongation factor Spt5 is relatively well-understood, the interactions involving Cdc73 have not been fully elucidated. Using a site-specific protein cross-linking strategy in yeast cells, we identified direct interactions between Cdc73 and two components of the Pol II elongation complex, the elongation factor Spt6 and the largest subunit of Pol II. Both of these interactions require the tandem SH2 domain of Spt6. We also show that Cdc73 and Spt6 can interact in vitro and that rapid depletion of Spt6 dissociates Paf1 from chromatin, altering patterns of Paf1C-dependent histone modifications genome-wide. These results reveal interactions between Cdc73 and the Pol II elongation complex and identify Spt6 as a key factor contributing to the occupancy of Paf1C at active genes in Saccharomyces cerevisiae.


Subject(s)
Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae , Nuclear Proteins/metabolism , Peptide Elongation Factors/metabolism , RNA Polymerase II/genetics , RNA Polymerase II/metabolism , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Transcription Factors/metabolism , Transcription, Genetic , Transcriptional Elongation Factors/genetics , Transcriptional Elongation Factors/metabolism
4.
Proc Natl Acad Sci U S A ; 119(12): e2116251119, 2022 03 22.
Article in English | MEDLINE | ID: mdl-35290126

ABSTRACT

RNA modifications regulate a variety of cellular processes including DNA repair.The RNA methyltransferase TRDMT1 generates methyl-5-cytosine (m5C) on messen-ger RNA (mRNA) at DNA double-strand breaks (DSBs) in transcribed regions, pro-moting transcription-coupled homologous recombination (HR). Here, we identifiedthat Fragile X mental retardation protein (FMRP) promotes transcription-coupled HRvia its interaction with both the m5C writer TRDMT1 and the m5C eraser ten-eleventranslocation protein 1 (TET1). TRDMT1, FMRP, and TET1 function in a temporalorder at the transcriptionally active sites of DSBs. FMRP displays a higher affinity forDNA:RNA hybrids containing m5C-modified RNA than for hybrids without modifica-tion and facilitates demethylation of m5C by TET1 in vitro. Loss of either the chroma-tin- or RNA-binding domain of FMRP compromises demethylation of damage-inducedm5C in cells. Importantly, FMRP is required for R-loop resolving in cells. Due to unre-solved R-loop and m5C preventing completion of DSB repair, FMRP depletion or lowexpression leads to delayed repair of DSBs at transcriptionally active sites and sensitizescancer cells to radiation in a BRCA-independent manner. Together, ourfindings presentan m5C reader, FMRP, which acts as a coordinator between the m5C writer and eraserto promote mRNA-dependent repair and cell survival in cancer.


Subject(s)
Fragile X Mental Retardation Protein , Fragile X Syndrome , Cytosine , Demethylation , Fragile X Mental Retardation Protein/genetics , Fragile X Mental Retardation Protein/metabolism , Fragile X Syndrome/genetics , Homologous Recombination , Humans , Mixed Function Oxygenases/metabolism , Proto-Oncogene Proteins/metabolism , RNA/genetics , RNA/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism
5.
Angew Chem Int Ed Engl ; : e202415168, 2024 Aug 16.
Article in English | MEDLINE | ID: mdl-39150898

ABSTRACT

Deubiquitinase-targeting chimera (DUBTAC) is a promising technology for inducing targeted protein stabilization (TPS). Despite its therapeutic potential, very few proteins have been stabilized by DUBTACs to date. The limited applicability of this technology is likely due to the modest DUBTAC-induced protein stabilization effect, and the scarcity of effective deubiquitinase ligands that can be harnessed for DUBTAC development. Here, we report the discovery of MS7829 and MS8588, the first-in-class DUBTACs of cGAS, a key component of the cGAS-STING pathway. While these DUBTACs are based on a cGAS inhibitor, they effectively stabilized cGAS and activated the cGAS/STING/IRF3 signaling. To develop these cGAS DUBTACs, we optimized EN523, an OTUB1 covalent ligand, into an improved ligand, MS5105. We validated MS5105 by generating a MS5105-based CFTR DUBTAC, which was approximately 10-fold more effective in stabilizing the ΔF508-CFTR mutant protein than the previously reported EN523-based CFTR DUBTAC. Overall, this work advances the DUBTAC technology for TPS.

6.
Diabetes Metab Res Rev ; 39(2): e3592, 2023 02.
Article in English | MEDLINE | ID: mdl-36401613

ABSTRACT

AIMS: This study investigated insulinoma-associated-2 autoantibody (IA-2A) and zinc transporter 8 autoantibody (ZnT8A) distribution in patients with type 1 diabetes (T1D) and latent autoimmune diabetes (LAD) and the autoantibodies' association with clinical characteristics and HLA-DR-DQ genes. MATERIALS AND METHODS: This cross-sectional study recruited 17,536 patients with diabetes from 46 hospitals across China. A total of 189 patients with T1D and 58 patients with LAD with IA-2A positivity, 126 patients with T1D and 86 patients with LAD with ZnT8A positivity, and 231 patients with type 2 diabetes (T2D) were selected to evaluate islet autoantibodies, clinical phenotypes, and HLA-DR-DQ gene frequency. RESULTS: IA-2A was bimodally distributed in patients with T1D and LAD. Patients with low IA-2A titre LAD had lower fasting C-peptide (FCP) (p < 0.01), lower postprandial C-peptide (PCP) (p < 0.001), and higher haemoglobin A1c (HbA1c) levels (p < 0.05) than patients with T2D. Patients with high IA-2A titre LAD were younger than patients with low IA-2A titre LAD (p < 0.05). Patients with low IA-2A titre T1D had lower FCP (p < 0.01), lower PCP (p < 0.01), and higher HbA1c levels (p < 0.05) than patients with high IA-2A titre LAD. HLA-DR-DQ genetic analysis demonstrated that the frequency of susceptible HLA haplotypes was higher in IA-2A-positive patients (p < 0.001) than in patients with T2D. Patients with high ZnT8A titre LAD had lower FCP (p = 0.045), lower PCP (p = 0.023), and higher HbA1c levels (p = 0.009) and a higher frequency of total susceptible haplotypes (p < 0.001) than patients with low ZnT8A titre LAD. CONCLUSIONS: IA-2A in patients with T1D and LAD was bimodally distributed, and the presence of IA-2A could demonstrate partial LAD clinical characteristics. ZnT8A titre had a certain predictive value for islet functions in patients with LAD.


Subject(s)
Cation Transport Proteins , Diabetes Mellitus, Type 1 , Diabetes Mellitus, Type 2 , Glucose Intolerance , Insulinoma , Pancreatic Neoplasms , Humans , Diabetes Mellitus, Type 1/genetics , Zinc Transporter 8 , Autoantibodies , Cross-Sectional Studies , C-Peptide , Glycated Hemoglobin , Cation Transport Proteins/genetics , HLA-DR Antigens , Glutamate Decarboxylase
7.
J Am Chem Soc ; 144(24): 10809-10816, 2022 06 22.
Article in English | MEDLINE | ID: mdl-35574633

ABSTRACT

Fluorosubstituted tryptophans serve as valuable probes for fluorescence and nuclear magnetic resonance (NMR) studies of proteins. Here, we describe an unusual photoreactivity introduced by replacing the single tryptophan in cyclophilin A with 7-fluoro-tryptophan. UV exposure at 282 nm defluorinates 7-fluoro-tryptophan and crosslinks it to a nearby phenylalanine, generating a bright fluorophore. The crosslink-containing fluorescent protein possesses a large quantum yield of ∼0.40 with a fluorescence lifetime of 2.38 ns. The chemical nature of the crosslink and the three-dimensional protein structure were determined by mass spectrometry and NMR spectroscopy. To the best of our knowledge, this is the first report of a Phe-Trp crosslink in a protein. Our finding may break new ground for developing novel fluorescence probes and for devising new strategies to exploit aromatic crosslinks in proteins.


Subject(s)
Phenylalanine , Tryptophan , Phenylalanine/chemistry , Spectrometry, Fluorescence , Tryptophan/chemistry
8.
Diabetes Obes Metab ; 23(6): 1282-1291, 2021 06.
Article in English | MEDLINE | ID: mdl-33528883

ABSTRACT

AIM: To investigate the frequency, clinical phenotype, inflammatory cytokine levels and genetics of glutamic acid decarboxylase autoantibody (GADA)-positive phenotypic youth-onset type 2 diabetes. MATERIALS AND METHODS: This nationwide, multicentre, cross-sectional study included 5324 newly diagnosed subjects with phenotypic type 2 diabetes aged 15 years or older enrolled in the LADA China study. GADA was screened in 248 subjects with youth-onset type 2 diabetes aged 15-29 years. Subjects who presented as GADA-positive were defined as having latent autoimmune diabetes in youth (LADY). We added subjects with LADY, type 1 diabetes, type 2 diabetes and controls from the Diabetes Center of Central South University, and measured serum concentrations of interleukin-6, lipocalin 2, high-sensitivity C-reactive protein, adiponectin and human leukocyte antigen (HLA) genotyping in subjects with LADY, age- and sex-matched GADA-negative type 2 diabetes, type 1 diabetes and controls. RESULTS: Twenty-nine of the 248 subjects (11.7%) were GADA positive. Compared with subjects with type 2 diabetes, subjects with LADY were less probable to have metabolic syndrome (27.6% vs. 59.4%; p = .001). The fasting C-peptide levels tended to be lower in subjects with LADY than in subjects with type 2 diabetes, but the difference was not statistically significant (LADY vs. type 2 diabetes: 0.21 [0.17-0.64] vs. 0.47 [0.29-0.77] nmol/L; p = .11). The cytokine levels of subjects with LADY were indistinguishable from subjects with type 1 diabetes, but subjects with LADY presented increased adiponectin levels compared with subjects with type 2 diabetes after adjusting for age, sex and body mass index (7.19 [4.05-11.66] vs. 3.42 [2.35-5.74] µg/mL; p < .05). The frequency of total susceptible HLA genotypes (DR3/3, -3/9 and -9/9) in subjects with LADY and type 1 diabetes were similarly higher than controls (LADY and type 1 diabetes vs. controls: 21.4% and 30.8% vs. 2.6%, respectively; p < .001). CONCLUSIONS: A high GADA positivity was observed in youth-onset type 2 diabetes subjects in China. As subjects with LADY had an increased susceptible HLA genetic load and different cytokine levels compared with subjects with type 2 diabetes, differentiating LADY from phenotypic type 2 diabetes subjects is important.


Subject(s)
Diabetes Mellitus, Type 1 , Diabetes Mellitus, Type 2 , Adolescent , Adult , Autoantibodies , China/epidemiology , Cross-Sectional Studies , Cytokines , Diabetes Mellitus, Type 1/epidemiology , Diabetes Mellitus, Type 1/genetics , Diabetes Mellitus, Type 2/epidemiology , Diabetes Mellitus, Type 2/genetics , Genetic Background , Humans , Young Adult
9.
J Autoimmun ; 111: 102452, 2020 07.
Article in English | MEDLINE | ID: mdl-32291137

ABSTRACT

The emergent outbreak of coronavirus disease 2019 (COVID-19) has caused a global pandemic. Acute respiratory distress syndrome (ARDS) and multiorgan dysfunction are among the leading causes of death in critically ill patients with COVID-19. The elevated inflammatory cytokines suggest that a cytokine storm, also known as cytokine release syndrome (CRS), may play a major role in the pathology of COVID-19. However, the efficacy of corticosteroids, commonly utilized antiinflammatory agents, to treat COVID-19-induced CRS is controversial. There is an urgent need for novel therapies to treat COVID-19-induced CRS. Here, we discuss the pathogenesis of severe acute respiratory syndrome (SARS)-induced CRS, compare the CRS in COVID-19 with that in SARS and Middle East respiratory syndrome (MERS), and summarize the existing therapies for CRS. We propose to utilize interleukin-6 (IL-6) blockade to manage COVID-19-induced CRS and discuss several factors that should be taken into consideration for its clinical application.


Subject(s)
Antibodies, Monoclonal, Humanized/therapeutic use , Coronavirus Infections/pathology , Cytokine Release Syndrome/therapy , Interleukin-6/antagonists & inhibitors , Pneumonia, Viral/pathology , Anti-Inflammatory Agents/therapeutic use , Betacoronavirus/pathogenicity , COVID-19 , Humans , Immunomodulation/drug effects , Interleukin-6/blood , Pandemics , SARS-CoV-2 , Severe Acute Respiratory Syndrome/pathology
10.
Diabetes Metab Res Rev ; 36(1): e3205, 2020 01.
Article in English | MEDLINE | ID: mdl-31318117

ABSTRACT

Latent autoimmune diabetes in adults (LADA) is the most common form of autoimmune diabetes diagnosed in adults. Similar to type 1 diabetes, the prevalence of LADA is impacted by ethnicity and geography. LADA is characterized by ß cell loss due to autoimmunity and insulin resistance and has highly heterogeneous clinical features, autoimmunity, and genetics in a glutamic acid decarboxylase antibody (GADA) titre-dependent manner, suggesting LADA is part of a continuum spectrum between type 1 and type 2 diabetes. Although LADA is the most frequent form of autoimmune diabetes diagnosed in adults, clinical trials involving LADA are scarce. Here we review the recent advancements in LADA epidemiology, clinical features, pathogenesis, and interventions. We also highlight the environmental factors that are thought to play an important role in addition to genetics in the pathogenesis of LADA. In the future, high-throughput molecular profiles might shed light on the nature of LADA among the wide spectrum of diabetes and offer new opportunities to identify novel LADA-specific biomarkers.


Subject(s)
Diabetes Mellitus, Type 1/epidemiology , Adult , Age of Onset , Autoantibodies/immunology , Autoimmunity/immunology , Diabetes Mellitus, Type 1/immunology , Humans
11.
Diabetes Metab Res Rev ; 36(2): e3209, 2020 02.
Article in English | MEDLINE | ID: mdl-31343818

ABSTRACT

BACKGROUND: There are abundant variations in the phenotypes and genetics of type 1 diabetes (T1D) patients across different races. This study aimed to assess differences between juvenile acute onset (JAO) and adult acute onset in Chinese T1D patients. METHODS: Seven hundred and fifty-one acute onset T1D patients were divided into two groups by the patient onset age as follows: the juvenile acute onset group (≤20 years, JAO group) and the adult acute onset group (>20 years, AAO group). Clinical characteristics, islet autoantibodies, and HLA class II haplotypes and genotypes were compared between these two groups. RESULTS: In comparison with AAO patients, JAO patients had significantly lower relative weights and lower triglyceride levels (P < .001, P < .01, respectively) but higher frequency of ketoacidosis (P < .001), higher daily insulin dosage (Pc  < .001), higher HbA1c (Pc  < .05), and higher HDL-cholesterol levels (Pc  < .01). The JAO group showed a higher prevalence of IA-2A, ZnT8A, and multiple autoantibodies than that in the AAO group (P < .001, P < .01, P < .001, respectively). Haplotypes for DRB1*0301-DQA1*03-DQB1*0201, DR3, DR4, DR9, and DR3/DR9 genotypes are highly associated with JAO susceptibility, whereas only DR3 and DR9 genotypes confer risk for AAO. In the JAO group but not the AAO group, DR3 is related to ZnT8A, and DR3/DR9 is related to IA-2A and multiple autoantibodies. CONCLUSIONS: These observations suggest that JAO patients markedly differ from AAO patients in their clinical manifestations and genetics in the Chinese T1D population. Notably, the DR3/DR9 genotype can facilitate the appearance of IA-2A or multiple autoantibodies in JAO patients.


Subject(s)
Asian People/genetics , Autoantibodies/immunology , Biomarkers/analysis , Diabetes Mellitus, Type 1/pathology , HLA Antigens/immunology , Haplotypes , Adolescent , Adult , Age Factors , Age of Onset , Aged , Alleles , Case-Control Studies , Child , Child, Preschool , Diabetes Mellitus, Type 1/genetics , Diabetes Mellitus, Type 1/immunology , Female , Follow-Up Studies , Genetic Predisposition to Disease , Genotype , Humans , Infant , Male , Middle Aged , Prognosis , Young Adult
12.
Diabetes Metab Res Rev ; 35(1): e3068, 2019 01.
Article in English | MEDLINE | ID: mdl-30160000

ABSTRACT

BACKGROUND: Latent autoimmune diabetes in adults (LADA) exhibits significant clinical heterogeneity, but the underlying causes remain unclear. The aim of this study was to investigate whether age of onset of LADA contributes to the observed clinical heterogeneity by comparing the clinical, metabolic, and immunogenetic characteristics between elderly and young LADA patients. METHODS: The cross-sectional study included a total of 579 patients with LADA which was further divided into elderly LADA (E-LADA) group (n = 135, age of onset ≥60 years) and young LADA (Y-LADA) group (n = 444, age of onset <60 years). Age-matched subjects with type 2 diabetes were served as control (E-T2D group, n = 622). Clinical characteristics, serum autoantibodies, and HLA-DQ haplotypes were compared among these groups. RESULTS: Compared with patients with Y-LADA, patients with E-LADA have better residual beta-cell function and higher level of insulin resistance (both P < .01), more metabolic syndrome characteristics, similar proportion of islet autoantibody positivity, and strikingly different HLA-DQ genetic background. In comparison with E-T2D patients, E-LADA patients tend to have similar metabolic syndrome prevalence, comparable C-peptide levels, and insulin resistance levels and share similar HLA-DQ genetic characteristics. CONCLUSIONS: Elderly LADA differs phenotypically and genetically from Y-LADA but has a clinical and genetic profile more similar to that of E-T2D. These distinct phenotypes could potentially help physicians better manage patients with E-LADA.


Subject(s)
Insulin Resistance/physiology , Latent Autoimmune Diabetes in Adults/diagnosis , Phenotype , Adult , Aged , Autoantibodies/blood , China , Cross-Sectional Studies , Female , Humans , Latent Autoimmune Diabetes in Adults/blood , Male , Middle Aged
13.
Diabetes Obes Metab ; 21(4): 893-902, 2019 04.
Article in English | MEDLINE | ID: mdl-30471182

ABSTRACT

AIMS: To investigate the prevalence of adult-onset autoimmune diabetes (ADM) and predisposition to autoimmune diseases by quantifying serum organ-specific autoantibodies in people with phenotype of type 2 diabetes (T2D). MATERIALS AND METHODS: We included a nationally representative sample of 46 239 adults aged ≥20 years from 14 provinces, of whom 4671 had diabetes, plus 1000 control subjects with normal glucose tolerance (NGT). Participants were screened centrally for autoantibodies to glutamic acid decarboxylase (GAD), islet antigen 2 (IA2) and zinc transporter isoform-8 (Znt8) and were defined as having ADM where positive for these antibodies. We then assayed thyroid peroxidase (TPO), tissue transglutaminase (tTG) and 21-hydroxylase (21-OH) autoantibodies in randomly selected participants with ADM and in age-matched, sex-matched and non-ADM controls with T2D plus controls with NGT. RESULTS: Post-normalization, the standardized prevalence rate of ADM was 6.0% (95% confidence interval [CI] 5.3-6.8) in initially non-insulin-requiring participants with ADM, corresponding to six million adults in China, in whom adjusted antibody positivity was: TPO autoantibodies 16.3% (95% CI 10.8-21.8), tTG autoantibodies 2.1% (95% CI 0.0-4.2), and 21-OH autoantibodies 1.8% (95% CI -0.2 to 3.8). Those participants with ADM who were GAD autoantibody-positive had high risk of TPO autoantibody positivity (odds ratio [OR] 2.39, P = 0.0031) and tTG autoantibody positivity (OR 6.98, P = 0.027), while those positive for IA2 autoantibodies had a high risk of tTG autoantibody positivity (OR 19.05, P = 0.001). CONCLUSIONS: A proportion of people with phenotype of T2D in China have ADM, with diabetes-associated autoantibodies, and may be at risk of developing other organ-specific autoimmune diseases; therefore, it may be clinically relevant to consider screening such Chinese populations.


Subject(s)
Autoantibodies/immunology , Diabetes Mellitus, Type 1/epidemiology , Diabetes Mellitus, Type 2/epidemiology , Adult , Aged , Autoantigens/immunology , Autoimmune Diseases/immunology , Case-Control Studies , China/epidemiology , Diabetes Mellitus, Type 1/immunology , Diabetes Mellitus, Type 1/metabolism , Diabetes Mellitus, Type 2/immunology , Diabetes Mellitus, Type 2/metabolism , Female , GTP-Binding Proteins/immunology , Glucose Tolerance Test , Glutamate Decarboxylase/immunology , Humans , Iodide Peroxidase/immunology , Iron-Binding Proteins/immunology , Male , Middle Aged , Phenotype , Prevalence , Protein Glutamine gamma Glutamyltransferase 2 , Risk , Steroid 21-Hydroxylase/immunology , Transglutaminases/immunology , Young Adult , Zinc Transporter 8/immunology
14.
Diabetes Metab Res Rev ; 32(6): 615-22, 2016 09.
Article in English | MEDLINE | ID: mdl-26787598

ABSTRACT

BACKGROUND: Latent autoimmune diabetes in adults (LADA) is a form of autoimmune diabetes with heterogeneous features. This study aimed to investigate the persistent status of glutamic acid decarboxylase antibody (GADA) in patients with LADA and its association with clinical characteristics. METHODS: This 3-year follow-up study enrolled 107 LADA and 40 type 2 diabetes mellitus (T2DM) patients from October 2005 to December 2013. GADA titer, epitopes, and clinical characteristics (including fasting C-peptide and HbA1c ) in LADA patients were assayed annually. The human leukocyte antigen DQ (HLA-DQ) genotypes were also analysed. The relationship between the persistence of GADA and the clinical characteristics was investigated in LADA patients. RESULTS: After 3-year follow-up, 36.5% (39/107) LADA patients remained GADA positive (persistently positive group), 19.6% (21/107) patients fluctuated positively and negatively (fluctuating group), and 43.9% (47/107) patients became GADA negative, among which 61.7% (29/47) seroconversions occurred within 6 months of follow-up (transiently positive group). The GADA persistently positive group possessed higher titer of GADA than transiently positive group and fluctuant group (all p = 0.000), higher reactivities to middle and C-terminal regions of GAD65 than those in transiently positive group (p = 0.001 and p = 0.000, respectively), and lower baseline fasting C-peptide level than T2DM patients and transiently positive group [415(31-1862) vs 620(220-1658) pmol/L, p = 0.014; and 415(31-1862) vs 705(64-1541) pmol/L, p = 0.017, respectively]. The GADA transiently positive group retained a higher HbA1c level when compared with T2DM patients (p = 0.023). In addition, the three LADA groups shared similar frequencies of HLA-DQ susceptible haplotypes that were higher as compared with T2DM. The GADA persistently positive group had a higher annual declining rate in fasting C-peptide than T2DM patients [-14%(-174-33%) vs -1%(-27-28%), p = 0.007]. CONCLUSION: The LADA patients with GADA transient positivity account for a large proportion, whose clinical characteristics and HLA-DQ haplotypes are different from those of T2DM. The patients with high titer GADA and reactivities to GADA65 middle and C-terminal regions showed a persistent GADA positivity, in which a worse baseline and accelerated decline of ß-cell function need early intervention in the practice. Copyright © 2016 John Wiley & Sons, Ltd.


Subject(s)
Autoantibodies/blood , Glucose Intolerance/immunology , Glutamate Decarboxylase/immunology , Latent Autoimmune Diabetes in Adults/immunology , Adult , Aged , Autoantibodies/immunology , Case-Control Studies , Female , Follow-Up Studies , Humans , Latent Autoimmune Diabetes in Adults/blood , Male , Middle Aged , Prognosis , Prospective Studies
15.
Zhonghua Yi Xue Za Zhi ; 95(20): 1563-7, 2015 May 26.
Article in Zh | MEDLINE | ID: mdl-26463602

ABSTRACT

OBJECTIVE: To observe the changes of islet function in ZnT8A positive and IA-2A positive patients so as to provide rationales for early intervention of LADA patients. METHODS: A total of 69 outpatients and inpatients with newly diagnosed LADA testing for GADA, IA-2A and ZnT8A by radioligand binding assay (RBA) were enrolled from 2005 to 2010, including 13 with double positive for GADA and ZnT8A (GADA+ZnT8A double positive group), 14 with double positive for GADA and IA-2A (GADA+IA-2A double positive group) and 42 with positive for GADA alone (GADA positive group), 40 type 2 diabetes mellitus (T2DM) patients negative for the above three autoantibodies were selected as control subjects (T2DM group). The baseline clinical data of all groups were compared and all patients were followed annually for 3 years. Fasting plasma glucose (FBG), HbA1c, fasting C-peptide (FCP), C-peptide 2 h after meal (2 h CP), triglyceride (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) were detected and compared among the above groups during the follow-ups. RESULTS: Compared to GADA positive group and T2DM group, the GADA+IA-2A double positive group had higher level of baseline FBG and HbA1c (10.5 ± 3.6) mmol/L and (9.3 ± 2.6) % vs (9.2 ± 2.0) mmol/L and (7.4 ± 2.0) %, P = 0.017, 0.011) and had lower level of baseline FCP (415 (68-934) vs 538 (224-1 146) and 643 (163-1 658) pmol/L, P = 0.015, 0.003). During the follow-ups, FCP decreased year-on-year in GADA+ZnT8A double positive, GADA+IA-2A double positive and GADA positive groups. The annual average rate of decrease in FCP was more rapid in GADA+ZnT8A double positive and GADA+IA-2A double positive groups than those in GADA positive alone and T2DM groups (all P < 0.01). CONCLUSIONS: The islet functions of newly diagnosed LADA patients with GADA and IA-2A positive are much worse. And emergence of ZnT8A and IA-2A may accelerate the failure of islet function with diabetic progress in LADA patients.


Subject(s)
Diabetes Mellitus, Type 1 , Islets of Langerhans , Adult , Autoantibodies , Diabetes Mellitus, Type 2 , Glucose Intolerance , Glutamate Decarboxylase , Humans
16.
J Immunol ; 188(10): 4747-58, 2012 May 15.
Article in English | MEDLINE | ID: mdl-22490442

ABSTRACT

B cells play a critical role in the pathogenesis of autoimmune diabetes. To investigate the mechanisms by which B cell depletion therapy attenuates islet ß cell loss and particularly to examine the effect of B cells on both diabetogenic and regulatory Ag-specific T cells, we generated a transgenic BDC2.5NOD mouse expressing human CD20 on B cells. This allowed us to deplete B cells for defined time periods and investigate the effect of B cell depletion on Ag-specific BDC2.5 T cells. We depleted B cells with anti-human CD20 Ab using a multiple injection protocol. We studied two time points, before and after B cell regeneration, to examine the effect on BDC2.5 T cell phenotype and functions that included antigenic response, cytokine profile, diabetogenicity, and suppressive function of regulatory T (T(reg)) cells. We found unexpectedly that B cell depletion induced transient aggressive behavior in BDC2.5 diabetogenic T cells and reduction in T(reg) cell number and function during the depletion period. However, after B cell reconstitution, we found that more regenerated B cells, particularly in the CD1d(-) fraction, expressed immune regulatory function. Our results suggest that the regenerated B cells are likely to be responsible for the therapeutic effect after B cell depletion. Our preclinical study also provides direct evidence that B cells regulate both pathogenic and T(reg) cell function, and this knowledge could explain the increased T cell responses to islet Ag after rituximab therapy in diabetic patients in a recent report and will be useful in design of future clinical protocols.


Subject(s)
Antigens, CD20/biosynthesis , B-Lymphocyte Subsets/immunology , Diabetes Complications/immunology , Epitopes, T-Lymphocyte/immunology , Lymphopenia/immunology , T-Lymphocyte Subsets/immunology , Animals , Antigens, CD20/genetics , Antigens, CD20/immunology , B-Lymphocyte Subsets/metabolism , B-Lymphocyte Subsets/pathology , Diabetes Complications/metabolism , Diabetes Complications/pathology , Humans , Lymphocyte Count , Lymphopenia/metabolism , Lymphopenia/pathology , Mice , Mice, Inbred NOD , Mice, Transgenic , Phenotype , T-Lymphocyte Subsets/metabolism , T-Lymphocyte Subsets/pathology
17.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 39(8): 818-24, 2014 Aug.
Article in Zh | MEDLINE | ID: mdl-25202951

ABSTRACT

OBJECTIVE: To establish a method for in vitro expansion of human natural CD4⁺CD25⁺ T regulatory cell (Treg) cells for clinical study and immunotherapy. METHODS: Human natural CD4⁺CD25⁺ Treg were isolated from peripheral blood monocyte cells (PBMCs) by magnetic activated cell sorting (MACS) and expanded by CD3/CD28 expansion beads, IL-2 and rapamycin. The number and the viability of the freshly isolated and expanded Treg were detemined by trypan blue staining. The phenotype and the purity of the freshly isolated and expanded Treg were analyzed by FACS. Treg suppression activity was assessed by mixed lymphocyte reaction (MLR) assay. RESULTS: Human natural Treg were expanded up to 2 000 folds after 3 weeks in culture, and the activity was more than 97%. The expanded Treg retained Treg phenotype as shown by their freshly isolated counterparts, and the purity of CD4⁺CD25⁺FoxP3⁺ Treg was (94.22 ± 2.12)%. The expanded Treg demonstrated a similar potent suppression of both proliferating auto- and allo- CD4⁺CD25⁻ effector T cells in vitro in a cell number-dependent manner. CONCLUSION: An in vitro expansion of human natural Treg was established to obtain large numbers of human Treg with highly suppressive phenotype and function, thereby providing a solution to the availability of sufficient human natural Treg in clinical study and immunotherapy.


Subject(s)
Cell Culture Techniques , T-Lymphocytes, Regulatory/cytology , Cell Separation , Cells, Cultured , Humans , Interleukin-2 , Leukocytes, Mononuclear , Lymphocyte Culture Test, Mixed
18.
Neuro Oncol ; 26(8): 1438-1452, 2024 Aug 05.
Article in English | MEDLINE | ID: mdl-38554116

ABSTRACT

BACKGROUND: The mesenchymal (MES) subtype of glioblastoma (GBM) is believed to be influenced by both cancer cell-intrinsic alterations and extrinsic cellular interactions, yet the underlying mechanisms remain unexplored. METHODS: Identification of microglial heterogeneity by bioinformatics analysis. Transwell migration, invasion assays, and tumor models were used to determine gene function and the role of small molecule inhibitors. RNA sequencing, chromatin immunoprecipitation, and dual-luciferase reporter assays were performed to explore the underlying regulatory mechanisms. RESULTS: We identified the inflammatory microglial subtype of tumor-associated microglia (TAM) and found that its specific gene integrin beta 2 (ITGB2) was highly expressed in TAM of MES GBM tissues. Mechanistically, the activation of ITGB2 in microglia promoted the interaction between the SH2 domain of STAT3 and the cytoplasmic domain of ITGB2, thereby stimulating the JAK1/STAT3/IL-6 signaling feedback to promote the MES transition of GBM cells. Additionally, microglia communicated with GBM cells through the interaction between the receptor ITGB2 on microglia and the ligand ICAM-1 on GBM cells, while an increased secretion of ICAM-1 was induced by the proinflammatory cytokine leukemia inhibitory factor (LIF). Further studies demonstrated that inhibition of cyclin-dependent kinase 7 substantially reduced the recruitment of SNW1 to the super-enhancer of LIF, resulting in transcriptional inhibition of LIF. We identified notoginsenoside R1 as a novel LIF inhibitor that exhibited synergistic effects in combination with temozolomide. CONCLUSIONS: Our research reveals that the epigenetic-mediated interaction of GBM cells with TAM drives the MES transition of GBM and provides a novel therapeutic avenue for patients with MES GBM.


Subject(s)
Brain Neoplasms , Glioblastoma , Leukemia Inhibitory Factor , Microglia , Signal Transduction , Glioblastoma/metabolism , Glioblastoma/pathology , Glioblastoma/genetics , Humans , Microglia/metabolism , Microglia/pathology , Mice , Brain Neoplasms/metabolism , Brain Neoplasms/pathology , Brain Neoplasms/genetics , Animals , Leukemia Inhibitory Factor/metabolism , Epithelial-Mesenchymal Transition , Gene Expression Regulation, Neoplastic , Cell Proliferation , Tumor Cells, Cultured , Xenograft Model Antitumor Assays , STAT3 Transcription Factor/metabolism , Cell Movement
19.
Cancer Biol Ther ; 25(1): 2338955, 2024 Dec 31.
Article in English | MEDLINE | ID: mdl-38680092

ABSTRACT

Gliomas are the most common type of primary brain tumor. Despite advances in treatment, it remains one of the most aggressive and deadly tumor of the central nervous system (CNS). Gliomas are characterized by high malignancy, heterogeneity, invasiveness, and high resistance to radiotherapy and chemotherapy. It is urgent to find potential new molecular targets for glioma. The TRPM channels consist of TRPM1-TPRM8 and play a role in many cellular functions, including proliferation, migration, invasion, angiogenesis, etc. More and more studies have shown that TRPM channels can be used as new therapeutic targets for glioma. In this review, we first introduce the structure, activation patterns, and physiological functions of TRPM channels. Additionally, the pathological mechanism of glioma mediated by TRPM2, 3, 7, and 8 and the related signaling pathways are described. Finally, we discuss the therapeutic potential of targeting TRPM for glioma.


•TRPM channels are widely expressed in the human body and play an important role in gliomas.• Abnormal expression of TRPM2, 3, 7, and 8 channels in gliomas is associated with disease severity and prognosis.•TRPM2, 3, 7, and 8 channels are effective targets in glioma.


Subject(s)
Brain Neoplasms , Glioma , TRPM Cation Channels , Humans , Glioma/metabolism , Glioma/pathology , Glioma/genetics , Glioma/drug therapy , TRPM Cation Channels/metabolism , TRPM Cation Channels/genetics , Brain Neoplasms/metabolism , Brain Neoplasms/pathology , Brain Neoplasms/drug therapy , Brain Neoplasms/genetics , Signal Transduction , Animals
20.
Diabetes Metab Res Rev ; 29(5): 363-8, 2013 Jul.
Article in English | MEDLINE | ID: mdl-23390067

ABSTRACT

BACKGROUND: The ZnT8A is an independent marker for diagnosis of type 1 diabetes mellitus. We investigated the distribution and clinical features of ZnT8A positive latent autoimmune diabetes in adult (LADA) patients to explore the potential diagnostic application. METHODS: A total of 3062 phenotypic T2DM patients were randomly selected from a national multicenter study - the LADA China Study. Radioligand binding assays were applied to detect the presence of ZnT8A, GADA and IA-2A. HbA1c , fasting C-peptide and serum lipid levels were followed up with ZnT8A positive patients. RESULTS: The positive prevalence of ZnT8A, GADA and IA-2A in phenotypic T2DM patients was 1.99% (61/3062), 6.43% (197/3062) and 1.96% (60/3062), respectively. The ZnT8A positivity was lower than that of GADA(x² = 74.8, p < 0.001) but was comparable with that of IA-2A (p > 0.05). The positivity of ZnT8A in IA-2A positive patients was higher than that in GADA positive patients (38.3% vs. 10.7%, x² = 24.8, p < 0.001). On the basis of GADA and IA-2A positivity, the ZnT8A assay enhanced the diagnostic prevalence of LADA from 7.58 to 8.62%. The LADA patients who were positive for ZnT8A had higher systolic blood pressure when compared with GADA positive cases (p = 0.049) and higher total cholesterol levels when compared with antibody-negative T2DM patients (p = 0.035). CONCLUSION: The detection of ZnT8A at the basis of GADA and IA-2A can improve diagnostic sensitivity of Chinese LADA.


Subject(s)
Autoantibodies/analysis , Autoimmune Diseases/diagnosis , Cation Transport Proteins/antagonists & inhibitors , Diabetes Mellitus, Type 2/diagnosis , Adult , Aged , Aged, 80 and over , Autoimmune Diseases/blood , Autoimmune Diseases/epidemiology , Autoimmune Diseases/immunology , Case-Control Studies , China/epidemiology , Cross-Sectional Studies , Diabetes Mellitus, Type 2/blood , Diabetes Mellitus, Type 2/epidemiology , Diabetes Mellitus, Type 2/immunology , Diagnosis, Differential , Follow-Up Studies , Glutamate Decarboxylase/antagonists & inhibitors , Humans , Male , Middle Aged , Prevalence , Radioligand Assay , Receptor-Like Protein Tyrosine Phosphatases, Class 8/antagonists & inhibitors , Sensitivity and Specificity , Zinc Transporter 8
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