Altered expression of imprinted genes in patients with cytogenetically normal­acute myeloid leukemia: Implications for leukemogenesis and survival outcomes.

Genomic imprinting, an epigenetic mechanism that regulates gene expression from parental chromosomes, holds substantial relevance in multiple cancers, including hematopoietic malignancies. In the present study, the expression of a panel of 16 human imprinted genes in bone marrow samples from 64 patients newly diagnosed with cytogenetically normal-acute myeloid leukemia (CN-AML) were examined alongside peripheral blood samples from 85 healthy subjects. The validated findings of the present study revealed significant upregulation of seven genes [COPI coat complex subunit gamma 2 (COPG2), H19 imprinted maternally expressed transcript (H19), insulin like growth factor 2 (IGF2), PEG3 antisense RNA 1 (PEG3-AS1), DNA primase subunit 2 (PRIM2), solute carrier family 22 member 3 SLC22A3 and Zinc finger protein 215 (ZNF215)] in patients with CN-AML (P<0.001). Notably, the expression level of H19 exhibited an inverse association with the survival duration of the patients (P=0.018), establishing it as a predictive marker for two- and five-year survival in patients with CN-AML. Kaplan-Meier analysis demonstrated that patients with lower H19 expression had superior two- and five-year survival rates compared with those with higher H19 expression. The results of the present study highlighted the association between loss of imprinting and leukemogenesis in CN-AML, underscoring the significance of H19 imprinting loss as a prognostic indicator for unfavorable two- and five-year survival in CN-AML patients.

Presume Why Probiotics May Not Provide Protection in Inflammatory Bowel Disease through an Azoxymethane and Dextran Sodium Sulfate Murine Model.

Recent studies have shown dysbiosis is associated with inflammatory bowel disease (IBD). However, trying to restore microbial diversity via fecal microbiota transplantation (FMT) or probiotic intervention fails to achieve clinical benefit in IBD patients. We performed a probiotic intervention on a simulated IBD murine model to clarify their relationship. IBD was simulated by the protocol of azoxymethane and dextran sodium sulfate (AOM/DSS) to set up a colitis and colitis-associated neoplasm model on BALB/c mice. A single probiotic intervention using Clostridium butyricum Miyairi (CBM) on AOM/DSS mice to clarify the role of probiotic in colitis, colitis-associated neoplasm, gut microbiota, and immune cytokines was performed. We found dysbiosis occurred in AOM/DSS mice. The CBM intervention on AOM/DSS mice failed to improve colitis and colitis-associated neoplasms but changed microbial composition and unexpectedly increased expression of proinflammatory IL-17A in rectal tissue. We hypothesized that the probiotic intervention caused dysbiosis. To clarify the result, we performed inverse FMT using feces from AOM/DSS mice to normal recipients to validate the pathogenic effect of dysbiosis from AOM/DSS mice and found mice on inverse FMT did develop colitis and colon neoplasms. We presumed the probiotic intervention to some extent caused dysbiosis as inverse FMT. The role of probiotics in IBD requires further elucidation.

Loss of ZNF215 imprinting is associated with poor five-year survival in patients with cytogenetically abnormal-acute myeloid leukemia.

Genomic imprinting is a form of epigenetic regulation and imprinted genes are silenced in a parental-specific manner. Imprinting is associated with various human diseases and cancers, but its roles in leukemogenesis remains elusive. In this study, the expression of a panel of 16 human imprinted genes was investigated using real-time quantitative polymerase chain reaction and 8 of them were further validated in 114 patients newly diagnosed with cytogenetically abnormal-acute myeloid leukemia (CA-AML) and 85 healthy subjects. Our results demonstrated upregulated expression of 8 imprinted genes (C15orf2, COPG2, H19, IGF2, PEG3-AS1, PRIM2, SLC22A3 and ZNF215) was observed in patients with CA-AML (p < 0.001). Patients' survival days were negatively correlated with the expression levels of H19 (p = 0.024), PGE3-AS1 (p = 0.038), and ZNF215 (p = 0.012). Multivariate logistic regression analysis further revealed the expression level ZNF215 can be used as a predictor for five-year survival for patients with CA-AML (p = 0.009) with a hazard ratio of 0.870 (95.0% confident interval: 0.784-0.965). Our results demonstrated that loss of imprinting of imprinted genes is critical for the leukemogenesis of AML under CA condition, and loss of ZNF215 imprinting is associated with poor five-year survival of patients with CA-AML.

Disrupted Expression of Circadian Clock Genes in Patients with Bronchial Asthma.

PURPOSE: Circadian clock is synchronized to the 24-hour day by the daily light-dark cycle and proper function of circadian rhythm is essential for many physiological processes. Disruption of circadian rhythm can affect disease processes and influence disease severity, treatment responses, and even survivorship. In this retrospective case-controlled study, we tried to explore whether expression of circadian clock genes was disturbed in patients with bronchial asthma. PATIENTS AND METHODS: We performed real-time quantitative reverse transcriptase-polymerase chain reactions to examine the expression of the nine core circadian clock genes (BMAL1, CK1ε, CLOCK, CRY1, CRY2, PER1, PER2, PER3, and TIM) in total leukocytes of peripheral blood collected at chest clinics from 120 patients with asthma and 60 health individuals. RESULTS: Expression levels of the nine circadian clock genes were significantly different between patients and healthy individuals, but not associated with the asthma control status. We also noted the difference of PER3 expression in asthmatic patients with and without nocturnal symptoms. In well-controlled asthmatics, expression of BMAL1, CK1ε, CLOCK, CRY1, CRY2, and PER1 was significantly lower in patients with nocturnal symptoms than those without nocturnal symptoms. However, in not well-controlled asthmatics, expression of only BMAL1, CK1ε, PER1, and PER2 was significantly different between patients with and without nocturnal symptoms. Binary logistic regression analysis selected BMAL1, CKIε, PER3, and TIM as independent factors for bronchial asthma and ROC curves showed the combined expression of these four genes enhanced the capability of predicting asthma (AUC=0.924; 95% CI=0.875-0.958; P<0.001). CONCLUSION: Our results showed altered expression of circadian clock genes in patients with bronchial asthma and down-regulated PER3 in patients with nocturnal symptoms. Altered expression of circadian clock genes was also observed in asthmatics with or without nocturnal symptoms in well- or not well-controlled subgroups. Combined expression of BMAL1, CKIε, PER3, and TIM could be a potential predictor for bronchial asthma.

Alternations of Circadian Clock Genes Expression and Oscillation in Obstructive Sleep Apnea.

Circadian misalignment plays an important role in disease processes and can affect disease severity, treatment outcomes, and even survivorship. In this study, we aim to investigate whether expression and daily oscillation patterns of core circadian clock genes were disturbed in patients with obstructive sleep apnea/hypopnea (OSA) syndrome. We performed real-time quantitative reverse transcriptase-polymerase chain reactions to examine the expression of the nine core circadian clock genes in leukocytes of peripheral blood collected at 12 AM, 6 AM, 12 PM, and 6 PM from 133 patients with OSA and 11 normal controls. Daily expression patterns of the nine circadian clock genes were observed in normal controls, but three of these genes (BMAL1, CLOCK, CRY2) were disrupted in patients with OSA. The expressions of eight circadian clock genes (except PER1) at midnight were significantly downregulated in patients with severe OSA. Binary logistic regression analysis selected CRY1 and PER3 as independent factors for severe OSA and showed that the combined expressions of CRY1 and PER3 enhanced the capability of predicting severe OSA (Odds ratio, 5.800; 95% CI, 1.978 to 17.004; p = 0.001). Our results show that combined expressions of CRY1 and PER3 at midnight could be a potential predictor for severe OSA.

Up-regulation of PER3 Expression Is Correlated with Better Clinical Outcome in Acute Leukemia.

BACKGROUND: Altered expression of circadian clock genes has been linked to various types of cancer. This study aimed to investigate whether these genes are also altered in acute myeloid leukemia (AML) and acute lymphoid leukemia (ALL). MATERIALS AND METHODS: The expression profiles of nine circadian clock genes of peripheral blood (PB) leukocytes from patients with newly-diagnosed AML (n=41), ALL (n=23) and healthy individuals (n=51) were investigated. RESULTS: In AML, the expression of period 1 (PER1), period 2 (PER2), period 3 (PER3), cryptochrome 1 (CRY1), cryptochrome 2 (CRY2), brain and muscle aryl hydrocarbon receptor nuclear translocator (ARNT)-like 1 (BMAL1), and timeless (TIM) was significantly down-regulated, while that of CK1ε was significantly up-regulated. In ALL, the expression of PER3 and CRY1 was significantly down-regulated, whereas those of CK1ε and TIM were significantly up-regulated. Recovery of PER3 expression was observed in both patients with AML and those with ALL who achieved remission but not in patients who relapsed after treatment. CONCLUSION: Circadian clock genes are altered in patients with acute leukemia and up-regulation of PER3 is correlated with a better clinical outcome.

Biphasic response of mitochondrial biogenesis to oxidative stress in visceral fat of diet-induced obesity mice.

AIMS: Studies in skeletal muscle demonstrate a strong association of mitochondrial dysfunction with insulin resistance (IR). However, there is still a paucity of knowledge regarding the alteration of mitochondria in adipose tissue (AT) in the pathogenesis of IR in obesity. We investigated the mitochondrial biogenesis in visceral fat (VF) and subcutaneous fat (SF) in C57BL/6J mice fed a high-fat high-sucrose diet for 12 months. RESULTS: Impairment of glucose tolerance and insulin sensitivity developed after 1 month of the diet and was associated with a prompt increase of VF. The VF adipocytes were larger than those in the SF and had increased expressions of HIF-1α and p-NFκB p65. However, the alteration of mitochondrial biogenesis did not occur in the early stage when increased intracellular reactive oxygen species (ROS), mitochondrial oxygen consumption rate, and mitochondrial ROS emerged at the 1st, 2nd and 2nd month, respectively. Until the 6th month, the VF had markedly increased mitochondrial DNA content and expression of PGC-1α, Tfam, ATP5A, and MnSOD. This increase of mitochondrial biogenesis was followed by a generalized decrease at the 12th month and the mitochondrial morphology altered markedly. In the late stage, although mitochondrial ROS decreased, the increased expression of 8-OHdG in VF continued. INNOVATION AND CONCLUSION: These data suggest that IR and ROS production occur before the biphasic changes of mitochondrial biogenesis in AT, and the VF plays a more crucial role.

Genotype and phenotype predictors of relapse of graves' disease after antithyroid drug withdrawal.

BACKGROUND: For patients with Graves' disease (GD), the primary goal of antithyroid drug therapy is to temporarily restore the patient to the euthyroid state and wait for a subsequent remission of the disease. This study sought to identify the predictive markers for the relapse of disease. METHODS: To do this, we studied 262 GD patients with long enough follow-up after drug withdrawal to determine treatment outcome. The patients were divided into three groups by time of relapse: early relapse group (n = 91) had an early relapse within 9 months, late relapse group (n = 65) had a relapse between 10 and 36 months, and long-term remission group (n = 106) were either still in remission after at least 3 years or relapsed after 3 years of drug withdrawal. We assessed the treatment outcome of 23 SNPs of costimulatory genes, phenotype and smoking habits. We used permutation to obtain p values for each SNP as an adjustment for multiple testing. Cox proportional hazards models was performed to assess the strength of association between the treatment outcome and clinical and laboratory variables. RESULTS: FOUR SNPS WERE SIGNIFICANTLY ASSOCIATED WITH DISEASE RELAPSE: rs231775 (OR 1.96, 95% CI 1.18-3.26) at CTLA-4 and rs745307 (OR 7.97, 95% CI 1.01-62.7), rs11569309 (OR 8.09, 95% CI 1.03-63.7), and rs3765457 (OR 2.60, 95% CI 1.08-6.28) at CD40. Combining risk alleles at CTLA-4 and CD40 improved the predictability of relapse. Using 3 years as the cutoff point for multivariate analysis, we found several independent predictors of disease relapse: number of risk alleles (HR 1.30, 95% CI 1.09-1.56), a large goiter size at the end of the treatment (HR 1.30, 95% CI 1.05-1.61), persistent TSH-binding inhibitory Ig (HR 1.64, 95% CI 1.15-2.35), and smoking habit (HR 1.60, 95% CI 1.05-2.42). CONCLUSION: Genetic polymorphism of costimulatory genes, smoking status, persistent goiter, and TSH-binding inhibitory Ig predict disease relapse.

Tissue-specific differences in mitochondrial DNA content in type 2 diabetes.

BACKGROUND: To investigate whether the effect of hyperglycemia on mitochondrial DNA (mtDNA) content is tissue-specific. METHOD: We compared the mtDNA contents in leg muscle, blood vessel, and peripheral leucocytes in seventeen patients with type 2 diabetes (T2DM) with those of seven controls. We measured 8-hydroxydeoxyguanosine (8-OHdG) expression in the muscles and thiobarbituric acid reactive substance (TBARS) in sera to evaluate oxidative stress. Immunohistochemical detection of peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC1-α), mitochondrial transcription factor A (Tfam), and apoptosis were performed in the muscle tissue. RESULTS: The mtDNA copy number was highest in muscle tissue, followed by blood vessel tissue, and lowest in leucocytes in both the diabetic and control subjects. The diabetic patients had less mtDNA content in the muscle than the controls (2.86±0.33 vs. 3.20±0.14, P=0.025), but more mtDNA content in the leucocytes (2.25±0.26 vs. 1.98±0.06, P=0.04). In both groups, there was a positive correlation between muscle tissue mtDNA content and the expression of 8-OHdG. Patients with T2DM had significantly increased 8-OHdG and TUNEL labeling index and non-significant increases in the expression of PGC1-α and Tfam. CONCLUSION: Oxidative stress stimulates mitochondrial biogenesis but induces a greater degree of apoptosis in diabetic patients, resulting in a decrease in muscle tissue mtDNA content.

Gly482Ser polymorphism in the peroxisome proliferator-activated receptor gamma coactivator-1alpha gene is associated with oxidative stress and abdominal obesity.

The objective of the study was to o investigate the relationship of the Gly482Ser (G482S) polymorphism in the peroxisome proliferator-activated receptor gamma coactivator-1alpha (PPARGC1A) gene and type 2 diabetes mellitus (T2DM), obesity, and oxidative status in Chinese adults. We enrolled 276 T2DM patients and 1049 nondiabetic subjects aged at least 35 years. The G482S variant was detected using polymerase chain reaction and restriction enzyme digestion. The levels of thiobarbituric acid reactive substance, an indicator of lipid peroxidation, were measured in plasma samples. The homeostasis model assessment-estimated insulin resistance (HOMA-IR) index was determined for nondiabetic subjects. P values were adjusted for age, sex, and body mass index by using a generalized linear model. In this series, there was no association between G482S polymorphism and T2DM and obesity (body mass index >25 kg/m(2)). However, the plasma fasting insulin levels and HOMA-IR indices were significantly higher in nondiabetic subjects harboring the variant (S/S) genotype than in those with the heterozygous (G/S) genotype. With regard to the effect of the different genotypes on body fat distribution, overweight nondiabetic subjects harboring the S/S or G/S genotype had a significantly higher waist-to-hip ratio than those with the wild-type (G/G) genotype. Furthermore, subjects with the S/S genotype had significantly higher serum thiobarbituric acid reactive substance levels than those with the G/G genotype; the diabetic group mainly contributed to this significant association (P < .001). In overweight, nondiabetic Chinese adults, G482S polymorphism in the PPARGC1A gene is associated with hyperinsulinemia, HOMA-IR indices, and abdominal obesity. Furthermore, in hyperglycemia, the S482 allele is related to increased oxidative stress.

Peripheral blood mitochondrial DNA content and dysregulation of glucose metabolism.

OBJECTIVE: The aim of this study was to examine the potential influence of insulin resistance (IR), hyperglycemia and oxidative stress on leucocytes mitochondrial DNA (mtDNA) content. RESEARCH DESIGN AND METHOD: One hundred twenty-five T2DM, 101 IFG and 70 normal subjects were enrolled in this study. The quantity of relative mtDNA content was measured by a real-time PCR and corrected by simultaneous measurement of the nuclear DNA. Parameters of lipid peroxidation, thiobarbituric acid reactive substance (TBARS), and total free thiols as antioxidative status were measured from serum samples. IR was assessed by homeostasis model assessment in the non-diabetic groups. Relationships among different variables were analyzed by general linear model correlation. RESULTS: In all subjects, after correcting for age, sex and BMI, there were progressive increases of leucocyte mtDNA copy number, TBARS, and total reduced thiols with progressive dysregulation of glucose metabolism (normal vs. IFG vs. T2DM). Furthermore, correlation between mtDNA content and glucose dysregulation persisted after sequential correction for age, sex, BMI and TBARS. The independent predictor of mtDNA content by regression analysis was hyperglycemia. In non-diabetic group, influence of family history of diabetes on mtDNA content turned to non-significant after correcting for fasting plasma glucose (FPG). Correlation study revealed that mtDNA content was correlated with FPG (P<0.001), but not IR. CONCLUSION: Our results indicate that hyperglycemia, not IR, is associated with an increase of leucocyte mtDNA copy number in cases of glucose dysregulation.

Fatty liver and chronic inflammation in Chinese adults.

OBJECTIVE: To investigate the significance of fatty liver as predictor of insulin resistance (IR) and chronic inflammation. RESEARCH DESIGN AND METHODS: This cross-sectional study included 450 adults of Han Chinese origin aged >or=35. Excluded were cases with hepatitis B or C, alcoholic liver disease, or currently using thiazolidinedione. The volunteers were screened for the presence of the components of metabolic syndrome (MtS). IR index was estimated by the homeostasis model assessment. The fatty liver index was evaluated by computed tomography, calculated as the liver/spleen (L/S) ratio arrived at by averaging Hounsfield values obtained for five 3-mm slices. Serum levels of adiponectin, C-reactive protein (CRP), leptin, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) were checked in 100 subjects with low-L/S ratio and 100 age- and sex-matched controls. RESULTS: Fatty liver index correlated with all MtS traits and IR index. The values of L/S ratios in subjects with 0, 1, 2, 3 and >or=4 traits of MtS were 1.25+/-0.13, 1.18+/-0.16, 1.12+/-0.21, 1.05+/-0.25 and 0.92+/-0.25, respectively (p<0.001). In our stepwise regression analysis to compare the L/S ratios to the conventional traits of MtS for association with adipokine dysregulation, we found L/S ratio to be independently associated with most of them: adiponectin (p<0.001), CRP (p<0.001), IL-6 (p=0.005) and TNF-alpha (p=0.014). CONCLUSION: In Chinese, fatty liver index correlated well with IR index and can be a better marker of chronic inflammation than the conventional components of MtS.

Cytotoxic T lymphocyte-associated molecule-4 gene polymorphism and hyperthyroid Graves' disease relapse after antithyroid drug withdrawal: a follow-up study.

CONTEXT: We previously showed an association between the exon1 +49 A/G single nucleotide polymorphism (SNP) and the relapse of Graves' disease (GD). The G allele was associated with early relapse. OBJECTIVE: In this follow-up study, we sought to replicate the result by genotyping nine additional polymorphisms and recruiting another 60 GD patients. DESIGN AND PARTICIPANTS: The GD patients were divided into three groups: recurred within 9 months, between 10-36 months, and more than 36 months. There were 65 patients with early recurrence, 55 with medium recurrence, and 88 with late recurrence. Although several SNPs were associated with recurrence, the most significant marker was still exon1 +49 A/G. Separate analysis of the genotypes for the 60 newly enrolled patients indicated that our present study was not biased by the previous samples. Once exon1 +49 A/G was included in the model to predict recurrence, other markers would not add more predictive information. Haplotype analysis did not show an additional value once exon1 +49 A/G was compulsorily included. RESULTS: Multivariate logistic regression analysis showed that GG genotype of exon1 +49 A/G SNP had an adjusted odds ratio of 2.2 (95% confidence interval, 1.1-4.4) compared with the combined group of GA plus AA. Other significant predictors were large goiter size at the end of the treatment and positive TSH-binding inhibitory Ig at the end of the treatment. CONCLUSIONS: This follow-up study confirms the usefulness of the exon1 +49 A/G SNP of the cytotoxic T lymphocyte-associated molecule-4 gene in predicting recurrence after cessation of treatment. There is no additional power by including other polymorphisms to predict recurrence.

Association of mitochondrial deoxyribonucleic acid 16189 variant (T->C transition) with metabolic syndrome in Chinese adults.

OBJECTIVE: A common variant in mitochondrial DNA (mtDNA) at bp 16189 (T-->C transition) has been associated with small birth size, adulthood hyperglycemia, and insulin resistance in Caucasians. In this study, we investigated whether mtDNA 16189 variant is associated with metabolic syndrome in Chinese subjects. METHODS: Six hundred fifteen Chinese adults, aged 40 yr or older, were recruited in this study. The 16189 variant of mtDNA was detected using PCR and restriction enzyme digestion. Metabolic syndrome was diagnosed on modified National Cholesterol Education Program Adult Treatment Panel III guidelines, using body mass index (BMI) instead of waist circumference. An association study was performed with chi2 test and logistic regression analysis. RESULTS: The prevalence of the 16189 variant was higher in patients with metabolic syndrome than in those without: 44% (125 of 284) vs. 33.2% (110 of 331) (P = 0.006). The association between this 16189 variant of mtDNA and metabolic syndrome (P = 0.021) remained significant even after correcting for age and BMI. As to the individual traits, the prevalence of fasting plasma glucose of at least 110 mg/dl (> or =6.1 mmol/liter) [(51.5% (121 of 235) vs. 42.1% (160 of 380); P = 0.023], type 2 diabetes mellitus [48.1% (113 of 235) vs. 39.2% (149 of 380); P = 0.031], and hypertriglyceridemia [44.3% (104 of 235) vs. 35.8% (136 of 380); P = 0.037] were significantly higher in subjects harboring the 16189 variant of mtDNA than those with the wild type. However, the prevalence of hypertension [53.2% (125 of 235) vs. 47.6% (181 of 380); P = 0.180], BMI greater than 25 kg/m2 [48.5% (114 of 235) vs. 43.9% (167 of 380); P = 0.270], and low high-density lipoprotein cholesterol [61.3% (144 of 235) vs. 54.7% (208 of 380); P = 0.111] did not reach a significant difference between the two groups. Furthermore, there was a trend of increasing frequency of occurrence of the 16189 variant in individuals having an increasing number of components of metabolic syndrome (Ptrend < 0.005). CONCLUSION: Our data strongly suggest that mtDNA 16189 variant underlies susceptibility to metabolic syndrome in the Chinese population.

Cytotoxic T lymphocyte-associated molecule-4 polymorphism and relapse of Graves' hyperthyroidism after antithyroid withdrawal.

We studied the A/G single nucleotide polymorphism (SNP) at position 49 in exon 1 of the cytotoxic T lymphocyte-associated molecule-4 gene in 148 Chinese Graves' disease (GD) patients and 171 controls. Our primary aim was to test for the association of this SNP with the relapse of the hyperthyroidism after antithyroid withdrawal. Our secondary aim was to investigate the relationship between GD patients and controls according to the SNP genotypes. All GD patients were divided into the following three groups according to the time of relapse after drug discontinuation: group 1, early relapse within 9 months; group 2, relapse between 10 and 36 months; and group 3, relapse 3 or more years after discontinuation of treatment. There was a significant difference of genotype frequencies (P < 0.001) and allele frequencies (P < 0.001) among the three groups of patients. The frequency of the G/G genotype decreased from 79% to 64% and 39% in groups 1, 2, and 3, respectively. Compared with controls, a strong association (P < 0.001) of G allele was found for group 1, and moderate significance (P = 0.04) was found for group 2, but no association (P = 0.33) was found for group 3. At the end of treatment, the percentage of patients with persistent TSH-receptor antibody was statistically different (A/A, 9.0%; A/G, 20.8%; G/G, 45.5%; P = 0.004). Using 3 yr as the cutoff point for multivariate logistic regression analysis, we found that the G/G genotype (adjusted odds ratio, 3.1 compared with A/G plus A/A; 95% confidence interval, 1.3-7.1), larger goiter size at the end of treatment, and positive TSH-receptor antibody at the end of treatment were independent risk factors of recurrence. We conclude that the A/G polymorphism of the cytotoxic T lymphocyte-associated molecule-4 gene affects the progress of GD. The G/G genotype is associated with poor outcome.