Juan Ouyang

Emerging tendency towards autoimmune process in major depressive patients: A novel insight from Th17 cells

Evidence indicates that there is an emerging tendency towards autoimmunity occuring in major depressive disorder (MDD). The aim of our study is to investigate the mechanism of autoimmune process in MDD from a novel insight of Th17 cells, which have been identified as the significant activators of autoimmunity. We included 40 patients with MDD and 30 healthy control subjects. An indirect immunofluorescence test was used for the detection of serum antinuclear antibodies (ANA), and revealed that the patient group was positive more frequently than the control group. By flow cytometric analysis, depressed subjects revealed a significant increase in peripheral Th17 cell number, and an obvious decrease in T-reg cell number, showing an imbalance of Th17/Treg ratio compared to healthy controls. We also found a higher level of RORγt (retinoic acid-related orphan receptor-γt, the specific transcription factor of Th17 cell) mRNA expression in peripheral blood lymphocytes by RT-PCR and serum concentration of IL-17 by ELISA in patients. In conclusion, our study showed a potential role of Th17 cells in the autoimmune process in MDD patients, thus contributing to the existing evidence of autoimmune inclination in MDD.

Urine particles analysis: Performance evaluation of Sysmex UF-1000i and comparison among urine flow cytometer, dipstick, and visual microscopic examination

Abstract Introduction. Our objective was to evaluate a newly invented urine flow cytometer, and select an optimal strategy for urinalysis in clinical practice. Methods. The performance of UF-1000i was evaluated in both control material and patient samples. A total of 1631 specimens were collected and analysed by visual microscopy examination (VME), UF-1000i flow cytometer (Sysmex Medical Electronics Co, Kobe, Japan) and an automated dipstick reflectometer Clinitek Atlas (Bayer Corp, Elkhart, USA). Results. UF-1000i showed good imprecision performance for the main parameters in urine particles with CV values less than 20%. The results from UF-1000i correlated well with VME for erythrocytes (r = 0.96), leukocytes (r = 0.98), and epithelial cell (r = 0.84). The area under the receiver operating curve (AUC) was 0.879, 0.903, 0.783, and 0.817 respectively for erythrocytes, leukocytes, bacteria and CAST in UF-1000i. While in Clinitek Atlas, the AUC was 0.848, 0.803, 0.761, and 0.754 respectively. Sensitivity of combination of the two methods for screening remained at 98% as compared to VME alone, while reducing the visual review rate down to 40%. Conclusion. UF-1000i is capable of reproducible measurement of urine particles in the clinically relevant range and shows its advantage over Atlas. Combination of the two methods is an optimal strategy for urine sample screening.

Epigenetic programming of diverse glucocorticoid response and inflammatory/immune-mediated disease

Glucocorticoid plays a fundamental role in maintaining immune homeostasis. Resistance to glucocorticoids is a potential etiology of inflammatory/immune-mediated disease. Most of the glucocorticoid effects are mediated by glucocorticoid receptor (GR), which has a complicated promoter region with multiple promoters. Studies have found that the methylation pattern of GR promoter is highly individual, which may contribute to the diverse glucocorticoid responds. Early life is a critical time for epigenetic programming of the body in which methylation imprints are established. Here we propose a hypothesis that connects the adverse early life events and the development of inflammatory/immune-mediated disease through an epigenetic mechanism, the methylation of GR gene.

Nuclear HSP90 regulates the glucocorticoid responsiveness of PBMCs in patients with idiopathic nephrotic syndrome.

Resistance to glucocorticoid (GC) is a challenge for the treatment of patients with idiopathic nephrotic syndrome (INS). Most of the effects of GC are mediated by the GC receptor (GR). Heat shock protein 90 (HSP90) is an important molecular chaperone for the GR and is supposed to be the key factor in regulating GC effects. In a previous study, we found that both the expression and nuclear distribution of HSP90 were increased in GC resistant INS patients. The aim of this study is to explore how these phenomena contribute to GC resistance in INS patients. Healthy subjects and INS patients with different GC responses were recruited. The total HSP90 expression was determined by reverse transcription-PCR and flow cytometric analysis. Western blot analysis was used to evaluate the expression of nuclear HSP90. Co-immunoprecipitation and electrophoretic mobility gel shift assays were performed to explore the interaction between HSP90 and the GR in the nucleus as well as the DNA-binding activity of GR. We induced the upregulation of the expression of total HSP90 in PBMCs by treatment with interleukin-6 in vitro and found that the nuclear HSP90 level, the DNA-binding activity of the GR and the cell apoptotic responsiveness to GC remained unchanged. Furthermore, an increased nuclear HSP90 was demonstrated mainly by binding to GR in the nucleus, while the DNA-binding activity of the GR dramatically decreased in GC resistant INS patients. The present results suggest that the accumulation of HSP90 in the nucleus potentially hinders DNA-binding activity and transactivation, which may contribute to GC resistance in patients with INS.

Depression, another autoimmune disease from the view of autoantibodies

Immune dysregulation is very common in major depression (MD) patients, with these individuals incurring increased risk for development of chronic inflammatory disease or autoimmune disease. Meanwhile, depressive symptoms have been found at a high prevalence in autoimmune disease. This apparent convergence suggests they may share a common pathogenic factor, or a close interaction innate. Recent studies have found that autoantibodies play an important role in the pathogenesis of depression both in animal models and human. Here, we suggest that depression, in nature, can be regarded as autoimmune disease caused by various autoantibodies, which broadens our understanding of depression, and brings about new fields for research and clinical implications of the disease.

Cell Atavistic Transition: Paired Box 2 Re-Expression Occurs in Mature Tubular Epithelial Cells during Acute Kidney Injury and Is Regulated by Angiotensin II

The regeneration of tubular epithelial cells (TECs) after acute kidney injury (AKI) is crucial for the recovery of renal structure and function. The mechanism by which quiescent TECs re-obtain a potential to regenerate remains unknown. In this study, we observed a transient re-expression of embryonic gene Paired box 2 (Pax2) in adult rat TECs in vivo during ischemia-reperfusion induced AKI and most Pax2 positive TECs co-expressed kidney injury molecule-1 (KIM-1), a tubular injury marker. The re-expression of Pax2 was accompanied by increased levels of intrarenal Angiotensin II, which is a crucial injury factor of AKI. Furthermore, we also found a temporary re-expression of Pax2 in NRK-52E cells under the stimulation of Angiotensin II. This stimulatory effect could be blocked by PD123319 (Angiotensin II type 2 receptor (AT2R) inhibitor) and AG490 (Janus Kinase 2 (JAK2) inhibitor). As Pax2 is essential for the phenotypic conversion from mesenchymal stem cells to TECs during kidney development, we proposed that the re-expression of Pax2 in mature TECs may be an indicator of “atavistic” transition which mimics but reverses the processes of development of TECs. This could be proved by that a progenitor marker, CD24, was also found to be transiently expressed shortly after the expression of Pax2 in NRK-52E cells stimulated with Angiotensin II. The expression of CD24 was also suppressed by PD123319 and AG490. Moreover, knockdown of Pax2 by RNA interference could significantly reduce the expression of CD24 in NRK-52E cells stimulated with Angiotension II. Those findings suggest that mature TECs can trans-differentiate into progenitor-like cells by “atavistic transition”, which may participate in the recovery of tissue structure and Pax2 may play a pivotal role in this process. That might have important implications for further understanding of tubular regeneration after injury.

Diagnostic value of the dual-luciferase report assay for predicting response to glucocorticoid in children with acute lymphoblastic leukemia

ObjectiveResistance to glucocorticoid (GC) is a significant clinical problem in some cases of acute lymphoblastic leukemia (ALL). Current methods of assessing GC resistance are time consuming and have limited reproducibility; in this study, we sought to define a new method of evaluating GC sensitivity and resistance in vitro.MethodsBased on the mechanisms of GC resistance, we hypothesized that the dual-luciferase report (DLR) assay could reflect the transcription effects of GC downstream of the GC-glucocorticoid receptor signaling pathway, thereby allowing the evaluation of reactions to GC. Sixty-two patients with differential GC response were included in this study. The prednisone induction test was used to divide the children with ALL into two groups: GC sensitive (GCS) and GC resistant (GCR). DLR assay was later conducted on those patients to evaluate its value for diagnosis of the GC reactivity. Receiver operating characteristic curves were used to identify the optimal assay cutoff for identifying response to GC.ResultsUsing the DLR assay analysis, we found that GCR subjects showed significantly lower reporter/control ratios for luciferase, as compared with GCS subjects. The optimal cutoff value for GC response was 0.67, with sensitivity of 77.1% and specificity of 93.3%. The DLR assay results were consistent with prednisone induction test results. Further, the DLR assay was simpler, more sensitive, and less time-consuming than the prednisone induction test.ConclusionsOur study showed that the DLR assay is relatively fast, simple, and sensitive. Accordingly, it could be useful for detecting GC response in children with ALL.

Study on the Clinical Significance of JAK2V617F Allele Burden in Philadelphia Chromosome-Negative Myeloproliferative Neoplasm.

BACKGROUND It was discovered that the somatic mutation in JAK2 exon 14 (JAK2V617F) totally modified the understanding and diagnosis of Philadelphia-Negative myeloproliferative neoplasm (Ph-MPNs), including polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). Real-time quantitative PCR is the most widely used method for JAK2V617F detection in clinical laboratory. In this study, we aimed to evaluate the clinical significance of JAK2V617F allele burden in Ph-MPNs detected by real-time quantitative PCR. METHODS A total of 208 bone marrow samples were collected from patients suspected to have Ph-MPNs. Real-time quantitative PCR was performed on each sample to obtain the JAK2V617F allele burden. Clinical and laboratory data from these participants were also recorded for their first visit. RESULTS Out of 208 participants, 118 patients were confirmed with Ph-MPNs. JAK2V617F mutations were found in 59 patients in the PV group (86.8%), 31 patients in the ET group (70.5%). PV, PMF, and ET showed a significant difference in the distribution of JAK2V617F allele burden. In JAK2V617F positive patients, JAK2V617F allele burden was closely related with WBC counts, platelet counts, and hemoglobin concentration. CONCLUSIONS JAK2V617F allele burden is a useful marker in the diagnosis, discrimination, and evaluation of PhMPNs.

Prenatal Dexamethasone Exposure Increases the Susceptibility to Autoimmunity in Offspring Rats by Epigenetic Programing of Glucocorticoid Receptor

Objective. Prenatal glucocorticoids (GC) can induce long term effects on offspring health. However, reports and related studies regarding the prolonged effects of prenatal GC on the development of autoimmunity are limited. Here, we aimed to explore the immunological effects of dexamethasone (DEX) exposure on young adults and whether glucocorticoid receptor (GR) is involved in this process. Methods. Wistar rats were given DEX during pregnancy. Susceptibility to autoimmunity in offspring was assessed using experimental autoimmune encephalomyelitis (EAE) and adjuvant-induced arthritis (AIA) animal models. To reveal the possible mechanism, glucocorticoid response, GR expression, and methylation status were measured in peripheral blood mononuclear cells (PBMCs). Results. Our results showed that the DEX-treated rats had greater susceptibility to EAE (100% versus 62.5%, P < 0.05) and AIA (63.6% versus 0%, P < 0.05) than saline control group. Glucocorticoid response and GR expression were decreased in DEX rats. Significant difference was also found in the methylation levels of GR exon 1-10 to exon 1-11 region. Conclusions. Prenatal DEX administration increases the susceptibility to autoimmune diseases, which is potentially mediated by programming GR methylation status and glucocorticoid sensitivity.

MicroRNA-185-5p restores glucocorticoid sensitivity by suppressing the mammalian target of rapamycin complex (mTORC) signaling pathway to enhance glucocorticoid receptor autoregulation

Abstract Overexpression of microRNA-185-5p (miR-185-5p) in glucocorticoid (GC)-sensitive acute lymphoblastic leukemia (ALL) was identified using a microarray and reverse transcription polymerase chain reaction and was further confirmed in ALL cell lines. A reporter assay confirmed that the Rictor-one component of mammalian target of rapamycin complex 2 (mTORC2) is a target of miR-185-5p. Decreased mTORC activity was also confirmed in GC-sensitive patients. Overexpression of miR-185-5p significantly enhanced GC sensitivity in CEM-C1 cells (GC resistance) by increasing the rate of cell apoptosis and cycle arrest, and decreasing cell survival, accompanied by a decrease in mTORC activity and an increase in GC-induced glucocorticoid receptor (GR) expression. Rapamycin, an mTORC1 inhibitor, showed similar effects to miR-185-5p. These results demonstrated that miR-185-5p enhances GC sensitivity via suppression of mTORC activity by enhancing GR autoupregulation and that miR-185-5p is a potential target for the diagnosis and reversion of GC resistance.