Mapping choline metabolites in normal and transformed cells.

INTRODUCTION: Choline is an essential human nutrient that is particular important for proliferating cells, and altered choline metabolism has been associated with cancer transformation. Yet, the various metabolic fates of choline in proliferating cells have not been investigated systematically. OBJECTIVES: This study aims to map the metabolic products of choline in normal and cancerous proliferating cells. METHODS: We performed 13C-choline tracing followed by liquid chromatography-high resolution mass spectrometry (LC-HRMS) analysis of metabolic products in normal and in vitro-transformed (tumor-forming) epithelial cells, and also in tumor-derived cancer cell lines. Selected metabolites were quantified by internal standards. RESULTS: Untargeted analysis revealed 121 LCMS peaks that were 13C-labeled from choline, including various phospholipid species, but also previously unknown products such as monomethyl- and dimethyl-ethanolamines. Interestingly, we observed formation of betaine from choline specifically in tumor-derived cells. Expression of choline dehydrogenase (CHDH), which catalyzes the first step of betaine synthesis, correlated with betaine synthesis across the cell lines studied. RNAi silencing of CHDH did not affect cell proliferation, although we observed an increased fraction of G2M phase cells with some RNAi sequences, suggesting that CHDH and its product betaine may play a role in cell cycle progression. Betaine cell concentration was around 10 µM, arguing against an osmotic function, and was not used as a methyl donor. The function of betaine in these tumor-derived cells is presently unknown. CONCLUSION: This study identifies novel metabolites of choline in cancer and normal cell lines, and reveals altered choline metabolism in cancer cells.

Dietary choline and betaine intake, choline-metabolising genetic polymorphisms and breast cancer risk: a case-control study in China.

Choline and betaine are essential nutrients involved in one-carbon metabolism and have been hypothesised to affect breast cancer risk. Functional polymorphisms in genes encoding choline-related one-carbon metabolism enzymes, including phosphatidylethanolamine N-methyltransferase (PEMT), choline dehydrogenase (CHDH) and betaine-homocysteine methyltransferase (BHMT), have important roles in choline metabolism and may thus interact with dietary choline and betaine intake to modify breast cancer risk. This study aimed to investigate the interactive effect of polymorphisms in PEMT, BHMT and CHDH genes with choline/betaine intake on breast cancer risk among Chinese women. This hospital-based case-control study consecutively recruited 570 cases with histologically confirmed breast cancer and 576 age-matched (5-year interval) controls. Choline and betaine intakes were assessed by a validated FFQ, and genotyping was conducted for PEMT rs7946, CHDH rs9001 and BHMT rs3733890. OR and 95 % CI were estimated using unconditional logistic regression. Compared with the highest quartile of choline intake, the lowest intake quartile showed a significant increased risk of breast cancer. The SNP PEMT rs7946, CHDH rs9001 and BHMT rs3733890 had no overall association with breast cancer, but a significant risk reduction was observed among postmenopausal women with AA genotype of BHMT rs3733890 (OR 0·49; 95 % CI 0·25, 0·98). Significant interactions were observed between choline intake and SNP PEMT rs7946 (P interaction=0·029) and BHMT rs3733890 (P interaction=0·006) in relation to breast cancer risk. Our results suggest that SNP PEMT rs7946 and BHMT rs3733890 may interact with choline intake on breast cancer risk.

LncRNA HOXA10-AS promotes the progression of esophageal carcinoma by regulating the expression of HOXA10.

Esophageal cancer (EC) remains a primary cause of cancer-associated fatality worldwide and is characterized by poor prognosis. HOXA10-AS is reported to be relevant with the development of different human cancers. However, its role and regulatory mechanism in EC are still obscure. Our study targeted at investigating the functional and mechanical roles of HOXA10-AS in EC. We confirmed by RT-qPCR that HOXA10-AS presented a remarkably high expression in EC cells. Functional experiments demonstrated that knocking down HOXA10-AS weakened proliferation, invasion and migration in vitro and impeded tumorigenesis in vivo. Further, we found that HOXA10-AS positively regulated its neighbor gene HOXA10 and influenced EC cell biological activities depending on HOXA10. Mechanistically, we showed that HOXA10-AS combined with FMR1 to target and stabilize HOXA10 mRNA. Moreover, HOXA10 served as a transcriptional factor to stimulate the transcription of its target gene CHDH. Finally, rescue assays confirmed that HOXA10 influenced EC cell growth through modulating CHDH. In conclusion, our study first determines the function of HOXA10-AS in EC and demonstrates its mechanism relating to HOXA10/CHDH, suggesting HOXA10-AS as a potential novel target for EC treatment. [Figure: see text].

Osmotic Contribution of Synthesized Betaine by Choline Dehydrogenase Using In Vivo and In Vitro Models of Post-traumatic Syringomyelia.

Introduction: Syringomyelia (SM) is a debilitating spinal cord disorder in which a cyst, or syrinx, forms in the spinal cord parenchyma due to congenital and acquired causes. Over time syrinxes expand and elongate, which leads to compressing the neural tissues and a mild to severe range of symptoms. In prior omics studies, significant upregulation of betaine and its synthesis enzyme choline dehydrogenase (CHDH) were reported during syrinx formation/expansion in SM injured spinal cords, but the role of betaine regulation in SM etiology remains unclear. Considering betaine's known osmoprotectant role in biological systems, along with antioxidant and methyl donor activities, this study aimed to better understand osmotic contributions of synthesized betaine by CHDH in response to SM injuries in the spinal cord. Methods: A post-traumatic SM (PTSM) rat model and in vitro cellular models using rat astrocytes and HepG2 liver cells were utilized to investigate the role of betaine synthesis by CHDH. Additionally, the osmotic contributions of betaine were evaluated using a combination of experimental as well as simulation approaches. Results: In the PTSM injured spinal cord CHDH expression was observed in cells surrounding syrinxes. We next found that rat astrocytes and HepG2 cells were capable of synthesizing betaine via CHDH under osmotic stress in vitro to maintain osmoregulation. Finally, our experimental and simulation approaches showed that betaine was capable of directly increasing meaningful osmotic pressure. Conclusions: The findings from this study demonstrate new evidence that CHDH activity in the spinal cord provides locally synthesized betaine for osmoregulation in SM pathophysiology. Supplementary Information: The online version of this article contains supplementary material available 10.1007/s12195-022-00749-5.

CHDH, a key mitochondrial enzyme, plays a diagnostic role in metabolic disorders diseases and tumor progression.

Human choline dehydrogenase (CHDH) is a transmembrane protein located in mitochondria. CHDH has been shown to be one of the important catalytic enzymes that catalyze the oxidation of choline to betaine and is involved in mitochondrial autophagy after mitochondrial damage. In recent years, an increasing number of studies have focused on CHDH and found a close association with the pathogenesis of various diseases, including tumor prognosis. Here we summarized the genomic localization, protein structure and basic functions of CHDH and discuss the progress of CHDH research in metabolic disorders and other diseases. Moreover, we described the regulatory role of CHDH on the progression of different types of malignant tumors. In addition, major pathogenic mechanisms of CHDH in multiple diseases may be associated with single nucleotide polymorphism (SNP). We look forward to providing new strategies and basis for clinical diagnosis and prognosis prediction of diseases by diagnosing SNP loci of CHDH genes. Our work evaluates the feasibility of CHDH as a molecular marker relevant to the diagnosis of some metabolic disorders diseases and tumors, which may provide new targets for the treatment of related diseases and tumors.

Nutritional Genomics in Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) is a common chronic condition associated with genetic and environmental factors in which fat abnormally accumulates in the liver. NAFLD is epidemiologically associated with obesity, type 2 diabetes, and dyslipidemia. Environmental factors, such as physical inactivity and an unbalanced diet, interact with genetic factors, such as epigenetic mechanisms and polymorphisms for the genesis and development of the condition. Different genetic polymorphisms seem to be involved in this context, including variants in PNPLA3, TM6SF2, PEMT, and CHDH genes, playing a role in the disease's susceptibility, development, and severity. From carbohydrate intake and weight loss to omega-3 supplementation and caloric restriction, different dietary and nutritional factors appear to be involved in controlling the onset and progression of NAFLD conditions influencing metabolism, gene, and protein expression. The polygenic risk score represents a sum of trait-associated alleles carried by an individual and seems to be associated with NAFLD outcomes depending on the dietary context. Understanding the exact extent to which lifestyle interventions and genetic predispositions can play a role in the prevention and management of NAFLD can be crucial for the establishment of a personalized and integrative approach to patients.

CHDH-PNPLA3 Gene-Gene Interactions Predict Insulin Resistance in Children with Obesity.

INTRODUCTION: Insulin resistance plays a major role in metabolic syndrome and is recognized as the most common risk factor for non-alcoholic fatty liver disease (NAFLD). Identifying predictors for insulin resistance could optimize screening and prevention. PURPOSE: To evaluate the contribution of multiple single nucleotide polymorphisms across genes related to NAFLD and choline metabolism, in predicting insulin resistance in children with obesity. METHODS: One hundred fifty-three children with obesity (73 girls), aged 7-18 years, were evaluated within the NutriGen Study (ClinicalTrials.gov-NCT02837367). Insulin resistance was defined by Homeostatic Model Assessment for insulin-resistance cut-offs that accommodated pubertal and gender differences. Anthropometric, metabolic, intake-related variables, and 55 single nucleotide polymorphisms related to NAFLD and choline metabolism were evaluated. Gene-gene interaction effects were assessed using Multiple Data Reduction Software. RESULTS: Sixty percent (93/153) of participants showed insulin resistance (58.7% of boys, 63% of girls). Children with insulin resistance presented significantly higher values for standardized body mass index, triglycerides, transaminases and plasma choline when compared to those without insulin resistance. Out of 52 single nucleotide polymorphisms analysed, the interaction between genotypes CHDH(rs12676) and PNPLA3(rs738409) predicted insulin resistance. The model presented a 6/10 cross-validation consistency and 0.58 testing accuracy. Plasma choline levels and alanine aminotransferase modulated the gene interaction effect, significantly improving the model. CONCLUSION: The interaction between genotypes in CHDH and PNPLA3 genes, modulated by choline and alanine aminotransferase levels, predicted insulin-resistance status in children with obesity. If replicated in larger cohorts, these findings could help identify metabolic risk in children with obesity.

Investigation of betaine as a novel psychotherapeutic for schizophrenia.

BACKGROUND: Betaine is known to act against various biological stresses and its levels were reported to be decreased in schizophrenia patients. We aimed to test the role of betaine in schizophrenia pathophysiology, and to evaluate its potential as a novel psychotherapeutic. METHODS: Using Chdh (a gene for betaine synthesis)-deficient mice and betaine-supplemented inbred mice, we assessed the role of betaine in psychiatric pathophysiology, and its potential as a novel psychotherapeutic, by leveraging metabolomics, behavioral-, transcriptomics and DNA methylation analyses. FINDINGS: The Chdh-deficient mice revealed remnants of psychiatric behaviors along with schizophrenia-related molecular perturbations in the brain. Betaine supplementation elicited genetic background-dependent improvement in cognitive performance, and suppressed methamphetamine (MAP)-induced behavioral sensitization. Furthermore, betaine rectified the altered antioxidative and proinflammatory responses induced by MAP and in vitro phencyclidine (PCP) treatments. Betaine also showed a prophylactic effect on behavioral abnormality induced by PCP. Notably, betaine levels were decreased in the postmortem brains from schizophrenia, and a coexisting elevated carbonyl stress, a form of oxidative stress, demarcated a subset of schizophrenia with "betaine deficit-oxidative stress pathology". We revealed the decrease of betaine levels in glyoxylase 1 (GLO1)-deficient hiPSCs, which shows elevated carbonyl stress, and the efficacy of betaine in alleviating it, thus supporting a causal link between betaine and oxidative stress conditions. Furthermore, a CHDH variant, rs35518479, was identified as a cis-expression quantitative trait locus (QTL) for CHDH expression in postmortem brains from schizophrenia, allowing genotype-based stratification of schizophrenia patients for betaine efficacy. INTERPRETATION: The present study revealed the role of betaine in psychiatric pathophysiology and underscores the potential benefit of betaine in a subset of schizophrenia. FUND: This study was supported by the Strategic Research Program for Brain Sciences from AMED (Japan Agency for Medical Research and Development) under Grant Numbers JP18dm0107083 and JP19dm0107083 (TY), JP18dm0107129 (MM), JP18dm0107086 (YK), JP18dm0107107 (HY), JP18dm0107104 (AK) and JP19dm0107119 (KH), by the Grant-in-Aid for Scientific Research on Innovative Areas from the MEXT under Grant Numbers JP18H05435 (TY), JP18H05433 (AH.-T), JP18H05428 (AH.-T and TY), and JP16H06277 (HY), and by JSPS KAKENHI under Grant Number JP17H01574 (TY). In addition, this study was supported by the Collaborative Research Project of Brain Research Institute, Niigata University under Grant Numbers 2018-2809 (YK) and RIKEN Epigenetics Presidential Fund (100214-201801063606-340120) (TY).

Identification of a Bipolar Disorder Vulnerable Gene CHDH at 3p21.1.

Genome-wide analysis (GWA) is an effective strategy to discover extreme effects surpassing genome-wide significant levels in studying complex disorders; however, when sample size is limited, the true effects may fail to achieve genome-wide significance. In such case, there may be authentic results among the pools of nominal candidates, and an alternative approach is to consider nominal candidates but are replicable across different samples. Here, we found that mRNA expression of the choline dehydrogenase gene (CHDH) was uniformly upregulated in the brains of bipolar disorder (BPD) patients compared with healthy controls across different studies. Follow-up genetic analyses of CHDH variants in multiple independent clinical datasets (including 11,564 cases and 17,686 controls) identified a risk SNP rs9836592 showing consistent associations with BPD (P meta = 5.72 × 10-4), and the risk allele indicated an increased CHDH expression in multiple neuronal tissues (lowest P = 6.70 × 10-16). These converging results may identify a nominal but true BPD susceptibility gene CHDH. Further exploratory analysis revealed suggestive associations of rs9836592 with childhood intelligence (P = 0.044) and educational attainment (P = 0.0039), a "proxy phenotype" of general cognitive abilities. Intriguingly, the CHDH gene is located at chromosome 3p21.1, a risk region implicated in previous BPD genome-wide association studies (GWAS), but CHDH is lying outside of the core GWAS linkage disequilibrium (LD) region, and our studied SNP rs9836592 is ∼1.2 Mb 3' downstream of the previous GWAS loci (e.g., rs2251219) with no LD between them; thus, the association observed here is unlikely a reflection of previous GWAS signals. In summary, our results imply that CHDH may play a previously unknown role in the etiology of BPD and also highlight the informative value of integrating gene expression and genetic code in advancing our understanding of its biological basis.