mRNA Expression Levels Of GATA-3 Research Articles

Maternal obesity exacerbates the responsiveness of offspring BALB/c mice to cow’s milk protein-induced food allergy

Food allergy has become a significant public health problem affecting a large number of people worldwide. Maternal obesity causes inflammation and alters the immune system of offspring, which may exacerbate their food allergy. The aim of this study was to determine whether offspring mice born to obese mothers would have more serve reactions to cow’s milk protein-induced food allergy, and further investigate the underlying mechanisms. Female offspring BALB/c mice of mothers with normal and high-fat diets were sensitized with β-lactoglobulin (BLG), respectively. Maternal obesity increased the serum immunoglobulin E and mouse mast cell protease levels, though did not have significant influence on anaphylactic symptom score, core temperature and diarrhea rate of offspring mice after BLG sensitization. Furthermore, maternal obesity led to a lower level of occludin mRNA expression in BLG -sensitized mice. The mice born to obese mothers exhibited increased mRNA expression levels of GATA-3, interleukin (IL)-4 and IL-10 in jejunum after BLG sensitization, indicating maternal obesity intensified Th2-type biased immune responses. In conclusion, maternal obesity exerted exacerbating effects on the responsiveness of their offspring to cow’s milk protein sensitization.

Frequent KRAS mutations in oncocytic papillary renal neoplasm with inverted nuclei

Papillary renal neoplasm with reverse polarity (PRNRP) is a newly documented rare tumour type. Its molecular pathological features have thus far been very little studied. There were 13 PRNRP cases including 3 The Cancer Genome Atlas (TCGA) cases and our 10 cases in this study. The 3 TCGA cases were found by a combined analysis of GATA3 mRNA expression levels and digital slides from the TCGA papillary renal cell carcinoma project. KRAS codon 12 mutations were identified in the three PRNRPs from TCGA. Of our 10 PRNRP cases, the mutations were also discovered using Sanger sequencing in seven (77.8%) of nine cases with available DNA, where KRAS p.G12V (n=3), p.G12D (n=2), p.G12R (n=1) and p.G12C (n=1) alterations were found. PRNRP shared similar gene expression profiles with renal distal tubules via an interprofile correlation analysis. Gene set enrichment analysis revealed that genes involved in 'KEGG aldosterone regulated sodium reabsorption' or 'hallmark apical surface' were enriched in PRNRP. Moreover, polarised immunostaining patterns for L1CAM and EMA in the distal tubule were maintained in PRNRP. These results imply that the tumour potentially originates from the distal tubule, especially from the cortical collecting duct, and probably retains its cell polarity, except for nuclear inversion. We therefore propose that oncocytic papillary renal neoplasm with inverted nuclei (OPRNIN) is a better name for this tumour type. OPRNIN is a kidney site-specific KRAS mutation neoplasm different from conventional papillary renal cell carcinoma.

Application of GATA-3 gene marker in the detection of hematologic disorders in children.

The aim of the present study was to investigate the use of GATA-3 markers in the detection of hematologic disorders in children. In total, 35 pediatric patients diagnosed with blood disease and treated in Henan Red Cross Blood Center from January 2014 to June 2015 were selected for the observation group. Another 32 healthy children were selected for the control group. The differences in the GATA-3 mRNA expression levels between the control and observation groups were detected via reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The differences in the GATA-3 protein expression levels were detected via enzyme-linked immunosorbent assay (ELISA) and western blot analysis. Compared with those in the healthy children, the mRNA expression levels of GATA-3 in patients with hematologic malignancies, acute lymphoblastic leukemia, myeloproliferative disorder, acute non-lymphocytic leukemia or thrombocytopenic purpura were significantly higher, and there were statistically significant differences between the groups (P<0.05). The results of ELISA showed that the GATA-3 protein expression levels in patients with hematologic malignancies (241.3±42.6 µg/l), acute lymphoblastic leukemia (196.3±21.6 µg/l), myeloproliferative disorder (284.2±45.1 µg/l), acute non-lymphocytic leukemia (269.3±31.4 µg/l) or thrombocytopenic purpura (272.1±39.1 µg/l) were significantly higher than those in healthy subjects (69.3±15.2 µg/l). The results of western blot analysis were consistent with those of ELISA. Based on our results, the expression levels of GATA-3 in healthy children and pediatric patients with blood diseases exhibit significant differences, and can be used as important markers for the clinical diagnosis of blood diseases in children.

Derp1-modified dendritic cells attenuate allergic inflammation by regulating the development of T helper type1(Th1)/Th2 cells and regulatory T cells in a murine model of allergic rhinitis

The CD4+CD25+Foxp3+ regulatory T cells (Tregs) are known to regulate Th2-induced allergic rhinitis (AR). In this study, we evaluated the efficacy of Derp1-modified dendritic cells (DCs) in AR immunotherapy. Derp1 was synthesized and transfected into DCs to generate Derp1-modified DCs. Phenotypes of Derp1-modified DCs were analyzed with flow cytometry using antibodies against DC markers CD11c, CD11b, CD59, CD103 and Toll-like receptor 1(TLR1). Four groups of subject mice were formed; the controls were treated with immature DCs, while the AR mice models were sensitized with Derp1(AR) and treated with DCs(DC-AR) or Derp1-modified DCs (Derp1DC-AR). The frequency of sneezing and scratching, eosinophil cell count, and Th1/Th2 ratio in the spleen were measured for all groups. The percentage of CD4+CD25+Foxp3+ Tregs in peripheral blood mononuclear cells was measured using flow cytometry; serum IgE, IgG1, and histamine were measured using enzyme-linked immunosorbent assay; expression levels of transcription factors T-bet, GATA3, Foxp3+ and IL-10 were analyzed using reverse transcription-polymerase chain reaction, and Western blot used in analyzed expression of Foxp3+ and IL-10 in nasal mucosa. Treatment with Derp1-modified DCs ameliorated the allergic response. The Derp1DC-AR group had significantly lower eosinophil cell count and the IgE, IgG1, and histamine levels than the AR and DC-AR groups, and higher mRNA levels of Th1 transcription factors T-bet, IL-10 and Foxp3 in nasal mucosa than DC-AR mice, but Th2 transcription factors GATA3 mRNA expression level has the opposite results. Furthermore, the Th1/Th2 ratio and percentage of CD4+CD25+Foxp3+ Tregs was significantly lower in the AR group (p<0.05), but higher in the Derp1DC-AR group than in the control group (p<0.01). Thus, the Derp1-modified DCs increased the percentage of CD4+CD25+Foxp3+Tregs and influenced the balance of Th1/Th2, showing an immunotherapeutic effect against AR.

Polymorphic Variant in GATA3 gene Is a Hallmark of PAR1-Deleted BCP-ALL and Associates with Poor Prognosis Among Pediatric Patients Treated with the BFM Backbone Protocols

Abstract Introduction Germline variant at rs3824662 in GATA3 gene was associated with early response to treatment as well as relapse risk in childhood BCP-ALL treated with COG protocols. This effect resulted from the strong link between the GATA3 polymorphism and the presence of somatic defects in leukemic cells including IKZF1 deletions, CRLF2 rearrangements and JAK2 gene mutations, which promote aggressive course of the disease. Aims The study aimed to evaluate an association between GATA3 gene polymorphism and clinical and biological features of pediatric BCP-ALL treated with the BFM backbone protocols. Methods Between November 2010 and June 2016, 957 consecutive children with newly diagnosed BCP-ALL were enrolled into the study. 822 patients treated according to ALL-IC BFM2009 protocol (n=594) and ALL-IC BFM2002 protocol (n=228) in 15 centers of the Polish Pediatric Leukemia/Lymphoma Study Group were enrolled into the study (median age 4.5 yrs, median follow-up time 2.5 yrs). Patients with BCR-ABL1 and MLL gene rearrangements were excluded from the analysis. The rs3824662 polymorphism of the GATA3 gene was genotyped using TaqMan probes. GATA3 mRNA expression level in leukemic cell was evaluated in BCP-ALL cases using qPCR with FAM-MGB probes (n=136). Targeted copy number screening of selected 23 loci was performed using the P335-B2 SALSA MLPA kit (MRC-Holland, Netherlands) in n=807 available DNA leukemia samples. MRD at day 15 and 33 was measured by flow cytometry with EuroFlow 8-color antibody panels. Results In the study group genotypes distribution withinrs3824662 of GATA3 was as follows: AA: n=44 (5.4%); CA: n=266 (32.4%); CC: n=512 (62.3%). Median MRD15 and MRD33 were 0.31% and 0.001% respectively. IKZF1 deletion were found in 18%, PAX5 in 20%, PAR1 in 12%, CDKN2A in 24%, CDKN2B 20%, BTG1 in 6.3%, ETV6 in 21%, EBF1 in 4% and RB1 in 6% of cases. Leukemic cells harbouring AA variant withinrs3824662 showed GATA3 mRNA expression 1.6 and 2.2 times higher compared to cells with CA and CC variants respectively, with the difference close to significant (ANOVA p=0.06). The presence of AA variant was not related to any gene deletion apart from microdeletions of the pseudoautosomal region PAR1 (Xp22 and Yp11) which occurred more frequently in AA carriers as compared to CA and CC carriers (11/41 vs. 34/242 vs. 52/468, p=0.013). We did not find any association of clinical features such as initial WBC, steroid response, sex and age at diagnosis among BCP-ALL patients with GATA3 genotype. However, AA carriers had a higher risk for MRD&gt;10% at day 15 OR (95%CI)=3.93 (1.37-11.23) as well as MRD&gt;0.01% at day 33, OR(95%CI)=2.95 (1.12-7.73) as compared CC carriers. Cox model of survival analysis was done for variables with univariate significance of p&lt;0.15 (risk group, sex). The results of Cox regression showedthe AA genotype remained associated with an increased risk of death regardless of risk group (assigned based on ALL-IC BFM09 protocol), HR(95%C)=2.95(1.12-7.73), P=0.028). Conclusions We showed that carriers of AA genotype in GATA3 are prone to develop PAR1 -deleted BCP-ALL. Moreover our study confirmed an association of GATA3 AA rs3824662 homozygosity with poor early response to treatment as well as risk of death among pediatric BCP-ALL patients treated with the BFM backbone protocols. Disclosures Madzio: National Science Centre: Research Funding.

Inflammation Enhances the Risks of Stroke and Death in Chronic Chagas Disease Patients.

Ischemic strokes have been implicated as a cause of death in Chagas disease patients. Inflammation has been recognized as a key component in all ischemic processes, including the intravascular events triggered by vessel interruption, brain damage and repair. In this study, we evaluated the association between inflammatory markers and the death risk (DR) and stroke risk (SR) of patients with different clinical forms of chronic Chagas disease. The mRNA expression levels of cytokines, transcription factors expressed in the adaptive immune response (Th1, Th2, Th9, Th17, Th22 and regulatory T cell), and iNOS were analyzed by real-time PCR in peripheral blood mononuclear cells of chagasic patients who exhibited the indeterminate, cardiac, digestive and cardiodigestive clinical forms of the disease, and the levels of these transcripts were correlated with the DR and SR. Cardiac patients exhibited lower mRNA expression levels of GATA-3, FoxP3, AHR, IL-4, IL-9, IL-10 and IL-22 but exhibited higher expression of IFN-γ and TNF-α compared with indeterminate patients. Digestive patients showed similar levels of GATA-3, IL-4 and IL-10 than indeterminate patients. Cardiodigestive patients exhibited higher levels of TNF-α compared with indeterminate and digestive patients. Furthermore, we demonstrated that patients with high DR and SR exhibited lower GATA-3, FoxP3, and IL-10 expression and higher IFN-γ, TNF-α and iNOS mRNA expression than patients with low DR and SR. A negative correlation was observed between Foxp3 and IL-10 mRNA expression and the DR and SR. Moreover, TNF-α and iNOS expression was positively correlated with DR and SR. Our data suggest that an inflammatory imbalance in chronic Chagas disease patients is associated with a high DR and SR. This study provides a better understanding of the stroke pathobiology in the general population and might aid the development of therapeutic strategies for controlling the morbidity and mortality of Chagas disease.

Changes in the expression of three markers in T lymphocytes of peripheral blood and immunoregulatory mechanisms of burned mice with sepsis at early stage

Changes in the expression of three markers in T lymphocytes of peripheral blood and immunoregulatory mechanisms of burned mice with sepsis at early stage

Characteristics of histone deacetylase 9 in peripheral blood of patients with bronchial asthma

To investigate the expression pattern of histone deacetylase 9 (HDAC9) in peripheral blood of asthmatics and its effect on immune cells (Th2, Th17, Tregs) involved in the pathogenesis of asthma. Forty-seven asthmatics from Ruijin Hospital were recruited and assigned to intermittent, mild and moderate-severe groups. Lung function test and Asthma Control Questionnaire were performed to evaluate asthma control and severity. Twenty healthy donors were enrolled as controls. GATA3, IL-4, and HDAC9 mRNA expression levels were measured by SYRB Green Real-time PCR. The cytokine IL-17-mainly produced by Th17 cells and TGF-β-mainly produced by Treg cells, were measured by ELISA. The GATA3 and IL-4 mRNA expression levels (28.12 ± 7.57 and 743.6 ± 312.8) were up-regulated in asthmatics as compared to the healthy controls [0.56 ± 0.22, 0.7 ± 0.8 (U = 16.00, 37.00, P < 0.01)]. The HDAC9 mRNA expression levels of intermittent, mild and moderate-severe groups were 3.20 ± 0.50, 89.6 ± 18.0, 323.0 ± 65.3, respectively, which were associated with the severity of disease (H = 11.32, P < 0.05). The level of IL-17 in asthmatic group was (83 ± 55) ng/L, which was up-regulated as compared to the healthy control [(34 ± 22) ng/L (U = 153.50, P < 0.01)]. The level of TGF-β was decreased in the asthmatic groups as compared to the healthy control, but the difference did not reach significance. HDAC9 mRNA expression level was positively correlated with GATA3 mRNA expression level (r = 0.482, P < 0.05), and also with IL-4 mRNA expression (r = 0.432, P < 0.05) and IL-17 (r = 0.538, P < 0.05), but negatively correlated with TGF-β (r = -0.417, P < 0.05). In patients with moderate-severe asthma, HDAC9 mRNA expression level was negatively correlated with FEV(1)% (r = -0.657, P < 0.05). HDAC9 mRNA expression was up-regulated in peripheral blood of asthmatics, which was not only associate with the Th2 master transcriptional factors GATA3, cytokine IL-4 mRNA, Th17 and Treg cell-related cytokines, but also with FEV(1)% in moderate-severe asthma.

GATA-3 is upregulated in peripheral blood mononuclear cells from patients with minimal change nephrotic syndrome

An imbalance of Th1 and Th2 cytokines has been reported in MCNS. Interleukin-13 (IL-13: Th2 cytokine) has been implicated in the pathogenesis of MCNS, but Th1/Th2 regulators such as T-bet (Th1-specific transcription factor) and GATA-3 (Th2-specific transcription factor) have not been examined. We isolated PBMC from 25 patients with MCNS during nephrosis and remission phases, from 17 nephrotic patients with membranous nephropathy (MN), and from 25 healthy subjects. We measured mRNA expression levels of T-bet, GATA-3, Stat5A (regulator of Th2 priming), IFN-gamma (Th1 cytokine), IL-2 (Th1 cytokine and activator of Stat5), IL-4 (Th2 cytokine), and IL-13 in PBMC, using real-time RT-PCR. GATA-3, Stat5A, and IL-13 mRNA expression levels were higher in the nephrotic MCNS group compared to the others. IL-2 mRNA expression levels were higher in nephrotic patients with MCNS and MN than in MCNS patients in remission and healthy controls. There were no differences in mRNA expression levels of T-bet, IFN-gamma, and IL-4 between MCNS and MN patients and healthy controls. This study is the first to reveal increased mRNA expression levels of GATA-3 and Stat5A in PBMC from MCNS patients in nephrosis. This study also supports recent findings suggesting the role of IL-13 in the development of MCNS. A predominant Th2 type of T cell activation may be involved in the pathogenesis of MCNS.

Preoperative GATA3 mRNA Expression in Peripheral Blood Mononuclear Cells is Up-Regulated in Patients With Postoperative Infection Following Hepatobiliary Pancreatic Surgery

Aggressive hepatobiliary pancreatic surgery has been associated with high complication rates. Correlations of Th1/Th2 balance and toll-like receptor (TLR) 2/4 expression with postoperative infection following surgery were prospectively evaluated. Plasma concentrations of interleukin (IL)-6, IL-10, soluble lymphocyte activation gene (sLAG)-3, and soluble CD30 were determined by enzyme-linked immunosorbent assay, and expression levels of T-bet, GATA-3, TLR2, and TLR4 mRNA in peripheral blood mononuclear cells were assayed by reverse transcription-polymerase chain reaction perioperatively in 56 consecutive patients who underwent hepatobiliary pancreatic surgery. Of the 56 patients, 30 patients had postoperative infection. Postoperative plasma levels of IL-6 and IL-10 were significantly higher in patients with postoperative infection than in those without infection (P < 0.05). Plasma soluble CD30 level and GATA-3 mRNA expression level were significantly higher preoperatively, and remained higher by postoperative d 7 in patients with postoperative infection (P < 0.05). Soluble lymphocyte activation gene levels were not significantly different between the two groups. T-bet mRNA expression level was significantly higher on postoperative d 3, 7, and 14 in patients with postoperative infection (P < 0.05). Preoperative expression levels of GATA-3 mRNA correlated significantly with those of TLR2 and TLR4 mRNA (P < 0.05). These results suggest that in patients with postoperative infection, Th1/Th2 balance shifts toward Th2 dominance preoperatively.

Profiles of type 1 and type 2 T cells in chronic idiopathic thrombocytopenic purpura

To explore the profiles of type 1 and type 2 T cells in chronic idiopathic thrombocytopenic purpura (ITP) patients. Samples of peripheral blood were collected from 30 chronic ITP patients, 8 males and 22 females, aged 34, 20 being in the active stage, and 10 in the remission stage, and 20 healthy persons, 7 males and 13 females, aged 31. Peripheral blood mononuclear cells (PBMCs) were isolated, cultured, and activated with PMA/ionomycin. Flow cytometry was used to measure the intracellular cytokines interferon (IFN)-gamma and interleukin (IL)-4 in the PBMCs so as to identify the Th1 cells (IFN-gamma(+) IL-4(+) CD4(+) cells), Th2 cells (IFN-gamma(-) IL-4(+) CD4(+) cells), Tc1 cells (IFN-gamma+ IL-4(-) CD8(+) cells), and Tc2 cells (IFN-gamma(-) IL-4(+) CD8(+) cells). The ratios of Th2/Tc2, Th1/Th2, and Tc1/Tc2 were calculated. Samples of spleen tissue were collected from 8 patients with chronic ITP, 3 males and 5 females, aged 30, and 6 patients with hereditary spherocytosis (HS), 3 males and 3 females, aged 35, who underwent splenectomy. Splenocytes were isolated, cultured, and activated with PMA/ionomycin. Real-time PCR was used to detect the mRNA expression of T-bet and GATA-3 in the PBMCs and splenocytes. The Th1/Th2 ratio of the patients in active stage was 25.62, significantly higher than those of the patients in remission stage (9.86) and the control (8.29, both P < 0.01), and the Tc1/Tc2 ratio of the patients in active stage was 30.23, significantly higher than those of the patients in remission stage (10.10) and the controls (12.58, both P < 0.01). The Th1/Th2 ratio of the splenocytes of the patients in active stage was 41.46, significantly higher than that of the controls (16.30, P < 0.01), and the Tc1/Tc2 ratio of the splenocytes of the active ITP patients was 35.80, not significantly higher than that of the controls (16.30, P = 0.082). The GATA-3 mRNA expression level of the PBMCs of the active ITP patients was one-fifth of that of the controls (P < 0.05) and the GATA-3 mRNA expression level of the splenocytes of the ITP patients was 0.34 of that of the HS patients (P < 0.05), however, there was no difference in the T-bet expression among the 3 groups. The T-bet/GATA-3 ratio of the PBMCs of the active ITP patients was 5.85 times that of the controls and the T-bet/GATA-3 ratio of the splenocytes of the active ITP patients was 2.68 times that of the controls (both P < 0.01). ITP is a T1 cells (Th1 and Tc1) predominant disease. The T-bet/GATA-3 ratio may provide a surrogate marker of T1/T2 cytokine balance. Shifting the cytokine patterns from T1 to T2 may be a potential immunotherapy for ITP.