C-terminally truncation is a prominent post-translational modification of human erythrocyte α-synuclein.

α-Synuclein is a protein related to synucleinopathies with high expression in the central nervous system and erythrocytes which are a major source of peripheral α-synuclein. Recent reports have suggested the presence of α-synuclein within extracellular vesicles derived from erythrocytes, potentially contributing to the pathogenesis of synucleinopathies. While Lewy bodies, intracellular inclusions containing aggregated α-synuclein, are prominently observed within the brain, their occurrence in peripheral neurons implies the dissemination of synucleinopathy pathology throughout the body via the propagation of α-synuclein. In this study, we found erythrocytes and circulating extracellular vesicles obtained from plasma contained α-synuclein, which was separated into four major forms using high-resolution clear native-PAGE and isoelectric focusing. Notably, erythrocyte α-synuclein was classified into full-length and C-terminal truncated forms, with truncation observed between Y133 and Q134 as determined by LC-MS/MS analysis. Our finding revealed that C-terminally truncated α-synuclein, which was previously reported to exist solely within the brain, was also present in erythrocytes and circulating extracellular vesicles obtained from plasma.

Identification of genotype-biochemical phenotype correlations associated with fructose 1,6-bisphosphatase deficiency.

Fructose-1,6-bisphosphatase (FBPase) deficiency, caused by an FBP1 mutation, is an autosomal recessive disorder characterized by hypoglycemic lactic acidosis. Due to the rarity of FBPase deficiency, the mechanism by which the mutations cause enzyme activity loss still remains unclear. Here we identify compound heterozygous missense mutations of FBP1, c.491G>A (p.G164D) and c.581T>C (p.F194S), in an adult patient with hypoglycemic lactic acidosis. The G164D and F194S FBP1 mutants exhibit decreased FBP1 protein expression and a loss of FBPase enzyme activity. The biochemical phenotypes of all previously reported FBP1 missense mutations in addition to G164D and F194S are classified into three functional categories. Type 1 mutations are located at pivotal residues in enzyme activity motifs and have no effects on protein expression. Type 2 mutations structurally cluster around the substrate binding pocket and are associated with decreased protein expression due to protein misfolding. Type 3 mutations are likely nonpathogenic. These findings demonstrate a key role of protein misfolding in mediating the pathogenesis of FBPase deficiency, particularly for Type 2 mutations. This study provides important insights that certain patients with Type 2 mutations may respond to chaperone molecules.

Extension of bacterial rDNA sequencing for simultaneous methylation detection and its application in microflora analysis.

Although polymerase chain reaction (PCR) amplification and sequencing of the bacterial 16S rDNA region has numerous scientific applications, it does not provide DNA methylation information. Herein, we propose a simple extension for bisulfite sequencing to investigate 5-methylcytosine residues in the bacterial 16S rDNA region from clinical isolates or flora. Multiple displacement amplification without DNA denaturation was used to preferentially pre-amplify single-stranded bacterial DNA after bisulfite conversion. Following the pre-amplification, the 16S rDNA region was analyzed using nested bisulfite PCR and sequencing, enabling the simultaneous identification of DNA methylation status and sequence data. We used this approach (termed sm16S rDNA PCR/sequencing) to identify novel methylation sites and a methyltransferase (M. MmnI) in Morganella morganii and different methylation motifs among Enterococcus faecalis strains from small volumes of clinical specimens. Further, our analysis suggested that M. MmnI may be correlated to erythromycin resistance. Thus, sm16S rDNA PCR/sequencing is a useful extension method for analyzing the DNA methylation of 16S rDNA regions in a microflora, providing additional information not provided by conventional PCR. Given the relationship between DNA methylation status and drug resistance in bacteria, we believe this technique can be effectively applied in clinical sample testing.

Post-translational modification of lysine residues in erythrocyte α-synuclein.

α-Synuclein is a protein linked to various synuclein-associated diseases ('synucleinopathies'), including Parkinson's disease, dementia with Lewy Bodies and multiple system atrophy, and is highly expressed in the central nervous system and in erythrocytes. Moreover, α-synuclein-containing erythrocyte-derived extracellular vesicles may be involved in the pathogenesis of synucleinopathies and their progression across the blood-brain barrier. Several post-translational modifications of α-synuclein have been reported in brain inclusions, including S129 phosphorylation, but fewer have been found in erythrocytes. In this study, we analysed the post-translational modifications of erythrocyte α-synuclein using liquid chromatography-mass spectrometry. We found that all lysine residues in the α-synuclein protein could be modified by acetylation, glycation, ubiquitination or SUMOylation but that phosphorylation, nitration and acylation were uncommon minor post-translational modifications in erythrocytes. Since the post-translational modification of lysine residues has been implicated in both membrane association and protein clearance, our findings provide new insight into how synucleinopathies may progress and suggest possible therapeutic strategies designed to target α-synuclein.

Proteogenomic landscape and clinical characterization of GH-producing pituitary adenomas/somatotroph pituitary neuroendocrine tumors.

The clinical characteristics of growth hormone (GH)-producing pituitary adenomas/somatotroph pituitary neuroendocrine tumors (GHomas/somatotroph PitNETs) vary across patients. In this study, we aimed to integrate the genetic alterations, protein expression profiles, transcriptomes, and clinical characteristics of GHomas/somatotroph PitNETs to identify molecules associated with acromegaly characteristics. Targeted capture sequencing and copy number analysis of 36 genes and nontargeted proteomics analysis were performed on fresh-frozen samples from 121 sporadic GHomas/somatotroph PitNETs. Targeted capture sequencing revealed GNAS as the only driver gene, as previously reported. Classification by consensus clustering using both RNA sequencing and proteomics revealed many similarities between the proteome and the transcriptome. Gene ontology analysis was performed for differentially expressed proteins between wild-type and mutant GNAS samples identified by nontargeted proteomics and involved in G protein-coupled receptor (GPCR) pathways. The results suggested that GNAS mutations impact endocrinological features in acromegaly through GPCR pathway induction. ATP2A2 and ARID5B correlated with the GH change rate in the octreotide loading test, and WWC3, SERINC1, and ZFAND3 correlated with the tumor volume change rate after somatostatin analog treatment. These results identified a biological connection between GNAS mutations and the clinical and biochemical characteristics of acromegaly, revealing molecules associated with acromegaly that may affect medical treatment efficacy.

MicroRNA-874 targets phosphomevalonate kinase and inhibits cancer cell growth via the mevalonate pathway.

The microRNA (miR) miR-874, a potential tumour suppressor, causes cell death via target gene suppression in various cancer types. Mevalonate pathway inhibition also causes cell death in breast cancer. However, the relationship between the mevalonate pathway and miR-874-induced apoptosis or its association with the tumour suppressor p53 has not been elucidated. We identified phosphomevalonate kinase (PMVK), a key mevalonate pathway enzyme, and sterol regulatory element-binding factor 2 (SREBF2), the master cholesterol biosynthesis regulator, as direct miR­874 targets. Next-generation sequencing analysis revealed a significant miR-874-mediated downregulation of PMVK and SREBF2 gene expression and p53 pathway enrichment. Luciferase reporter assays showed that miR-874 directly regulated PMVK and SREBF2. miR-874-induced apoptosis was p53 dependent, and single-cell RNA sequencing analysis demonstrated that miR-874 transfection resulted in apoptosis and p53 pathway activation. Downregulation of PMVK expression also caused cell cycle arrest and p53 pathway activation, which was rescued by geranylgeranyl pyrophosphate (GGPP) supplementation. Analysis of The Cancer Genome Atlas (TCGA) database indicated a negative correlation between miR-874 and PMVK expression and between miR-874 and SREBF2 expression. These findings suggest that miR-874 suppresses the mevalonate pathway by targeting SREBF2 and PMVK, resulting in GGPP depletion, which activates the p53 pathway and promotes cycle arrest or apoptosis.

Liver group 2 innate lymphoid cells regulate blood glucose levels through IL-13 signaling and suppression of gluconeogenesis.

The liver stores glycogen and releases glucose into the blood upon increased energy demand. Group 2 innate lymphoid cells (ILC2) in adipose and pancreatic tissues are known for their involvement in glucose homeostasis, but the metabolic contribution of liver ILC2s has not been studied in detail. Here we show that liver ILC2s are directly involved in the regulation of blood glucose levels. Mechanistically, interleukin (IL)-33 treatment induces IL-13 production in liver ILC2s, while directly suppressing gluconeogenesis in a specific Hnf4a/G6pc-high primary hepatocyte cluster via Stat3. These hepatocytes significantly interact with liver ILC2s via IL-13/IL-13 receptor signaling. The results of transcriptional complex analysis and GATA3-ChIP-seq, ATAC-seq, and scRNA-seq trajectory analyses establish a positive regulatory role for the transcription factor GATA3 in IL-13 production by liver ILC2s, while AP-1 family members are shown to suppress IL-13 release. Thus, we identify a regulatory role and molecular mechanism by which liver ILC2s contribute to glucose homeostasis.

Correction of Hypercortisolemia with an Improved Cognitive Function and Muscle Mass after Transsphenoidal Surgery in an Older Patient with Cushing's Disease.

Cushing's disease causes numerous metabolic disorders, cognitive decline, and sarcopenia, leading to deterioration of the general health in older individuals. Cushing's disease can be treated with transsphenoidal surgery, but thus far, surgery has often been avoided in older patients. We herein report an older woman with Cushing's disease whose cognitive impairment and sarcopenia improved after transsphenoidal surgery. Although cognitive impairment and sarcopenia in most older patients show resistance to treatment, our case indicates that normalization of the cortisol level by transsphenoidal surgery can be effective in improving the cognitive impairment and muscle mass loss caused by Cushing's disease.

Epigenetic modifications underlie the differential adipogenic potential of preadipocytes derived from human subcutaneous fat tissue.

Ceiling culture-derived preadipocytes (ccdPAs) and adipose-derived stem cells (ASCs) can be harvested from human subcutaneous fat tissue using the specific gravity method. Both cell types possess a similar spindle shape without lipid droplets. We previously reported that ccdPAs have a higher adipogenic potential than ASCs, even after a 7-wk culture. We performed a genome-wide epigenetic analysis to examine the mechanisms contributing to the adipogenic potential differences between ccdPAs and ASCs. Methylation analysis of cytosines followed by guanine (CpG) using a 450-K BeadChip was performed on human ccdPAs and ASCs isolated from three metabolically healthy females. Chromatin immunoprecipitation sequencing was performed to evaluate trimethylation at lysine 4 of histone 3 (H3K4me3). Unsupervised machine learning using t-distributed stochastic neighbor embedding to interpret 450,000-dimensional methylation assay data showed that the cells were divided into ASC and ccdPA groups. In Kyoto Encyclopedia of Genes and Genomes pathway analysis of 1,543 genes with differential promoter CpG methylation, the peroxisome proliferator-activated receptor (PPAR) and adipocytokine signaling pathways ranked in the top 10 pathways. In the PPARγ gene, H3K4me3 peak levels were higher in ccdPAs than in ASCs, whereas promoter CpG methylation levels were significantly lower in ccdPAs than in ASCs. Similar differences in promoter CpG methylation were also seen in the fatty acid-binding protein 4 and leptin genes. In conclusion, we analyzed the epigenetic status of adipogenesis-related genes as a potential mechanism underlying the differences in adipogenic differentiation capability between ASCs and ccdPAs.

Renal Injuries in Primary Aldosteronism: Quantitative Histopathological Analysis of 19 Patients With Primary Adosteronism.

[Figure: see text].

Position paper from the Japan Thyroid Association task force on the management of low-risk papillary thyroid microcarcinoma (T1aN0M0) in adults.

The incidence of thyroid carcinoma has been increasing worldwide. This is interpreted as an increase in the incidental detection of papillary thyroid microcarcinomas (PTMCs). However, mortality has not changed, suggesting overdiagnosis and overtreatment. Prospective clinical trials of active surveillance for low-risk PTMC (T1aN0M0) have been conducted in two Japanese institutions since the 1990s. Based on the favorable outcomes of these trials, active surveillance has been gradually adopted worldwide. A task force on the management of PTMC in adults organized by the Japan Thyroid Association therefore conducted a systematic review and has produced the present position paper based on the scientific evidence concerning active surveillance. This paper indicates evidence for the increased incidence of PTMC, favorable surgical outcomes for low-risk PTMC, recommended criteria for diagnosis using fine needle aspiration cytology, and evaluation of lymph node metastasis (LNM), extrathyroidal extension (ETE) and distant metastasis. Active surveillance has also been reported with a low incidence of disease progression and no subsequent recurrence or adverse events on survival if conversion surgery was performed at a slightly advanced stage. Active surveillance is a safe and valid strategy for PTMC, because it might preserve physical quality of life and reduce 10-year medical costs. However, some points should be noted when performing active surveillance. Immediate surgery is needed for PTMC showing high-risk features, such as clinical LNM, ETE or distant metastasis. Active surveillance should be performed under an appropriate medical team and should be continued for life.

TAS4464, a NEDD8-activating enzyme inhibitor, activates both intrinsic and extrinsic apoptotic pathways via c-Myc-mediated regulation in acute myeloid leukemia.

TAS4464, a potent, selective small molecule NEDD8-activating enzyme (NAE) inhibitor, leads to inactivation of cullin-RING E3 ubiquitin ligases (CRLs) and consequent accumulations of its substrate proteins. Here, we investigated the antitumor properties and action mechanism of TAS4464 in acute myeloid leukemia (AML). TAS4464 induced apoptotic cell death in various AML cell lines. TAS4464 treatments resulted in the activation of both the caspase-9-mediated intrinsic apoptotic pathway and caspase-8-mediated extrinsic apoptotic pathway in AML cells; combined treatment with inhibitors of these caspases markedly diminished TAS4464-induced apoptosis. In each apoptotic pathway, TAS4464 induced the mRNA transcription of the intrinsic proapoptotic factor NOXA and decreased that of the extrinsic antiapoptotic factor c-FLIP. RNA-sequencing analysis showed that the signaling pathway of the CRL substrate c-Myc was enriched after TAS4464 treatment. Chromatin immunoprecipitation (ChIP) assay revealed that TAS4464-induced c-Myc bound to the PMAIP1 (encoding NOXA) and CFLAR (encoding c-FLIP) promoter regions, and siRNA-mediated c-Myc knockdown neutralized both TAS4464-mediated NOXA induction and c-FLIP downregulation. TAS4464 activated both caspase-8 and caspase-9 along with an increase in NOXA and a decrease in c-FLIP, resulting in complete tumor remission in a human AML xenograft model. These findings suggest that NAE inhibition leads to anti-AML activity via a novel c-Myc-dependent apoptosis induction mechanism.

Two cases of symptomatic secondary hypophysitis due to Rathke's cleft cysts treated with glucocorticoids: long-term follow-up.

Rathke's cleft cyst (RCC) is a common incidental tumor in the hypothalamic-pituitary region. Some reports have shown that the clinical symptoms and endocrine functions of symptomatic RCCs are temporarily improved by glucocorticoid administration. However, it is still unknown whether glucocorticoid treatment is effective for symptomatic RCCs according to long-term observations. In this study, we describe the long-term clinical outcomes of two cases of glucocorticoid-treated biopsy-proven secondary hypophysitis caused by RCCs. We summarize the symptoms, imaging findings, and endocrine evaluations of two symptomatic RCC patients with concomitant hypophysitis before and after prednisolone treatment. In both evaluated cases, visual impairments and altered endocrine parameters were present due to chiasm and stalk compression; these outcomes improved after shrinkage of RCCs in response to prednisolone administration, and partial recovery of anterior pituitary hormone secretion was observed. However, in both cases, the deficits in anterior pituitary hormone secretion recurred, possibly due to persistent inflammatory infiltration in the RCCs and pituitary glands. After relapse of hypophysitis, anterior hormone secretion did not fully recover. In our cases of secondary hypophysitis caused by RCCs, prednisolone administration had an early effect of cyst shrinkage, followed by partial improvements in clinical symptoms and pituitary functions. However, long-term observation showed that prednisolone treatment did not contribute to complete improvement in anterior pituitary hormone dysfunction.

Adrenalectomy in Japanese patients with subclinical Cushing syndrome: 1-mg dexamethasone suppression test to predict the surgical benefit.

OBJECTIVES: To investigate whether the result of the 1-mg dexamethasone suppression test can predict the improvement of comorbidities after adrenalectomy in patients with subclinical Cushing syndrome. METHODS: This retrospective study included 117 subclinical Cushing syndrome patients who underwent adrenalectomy. The numbers of prescribed drugs for metabolic comorbidities and the clinical variables at diagnosis were compared with those at the follow up. Patients were classified into subgroups according to the result of the 1-mg dexamethasone suppression test. RESULTS: Significant improvements in blood pressure, serum cholesterol and body mass index were observed. Furthermore, a significant improvement in glycated hemoglobin was observed in patients with diabetes mellitus. These improvements led to a discontinuation or reduction of prescribed drugs after surgery. In addition, the greatest reduction of prescribed drugs was observed in patients whose serum cortisol levels were between 1.8 and 3.0 µg/dL after the 1-mg dexamethasone suppression test. CONCLUSIONS: The result of the 1-mg dexamethasone suppression test can be a useful factor predicting the improvement of comorbidities after adrenalectomy. Current data might give us a new insight into the decision-making for the treatment of subclinical Cushing syndrome.

131I-6ß-iodomethyl-19-norcholesterol adrenal scintigraphy as an alternative to adrenal venous sampling in differentiating aldosterone-producing adenoma from bilateral idiopathic hyperaldosteronism.

OBJECTIVE: To assess the correlation value between adrenal venous sampling (AVS) and I-6ß-iodomethyl-19-norcholesterol (NP-59) adrenal scintigraphy in differentiating aldosterone-producing adenoma (APA) from bilateral idiopathic hyperaldosteronism (BHA), and the use of NP-59 scintigraphy as an alternative to AVS. METHODS: Overall, 29 patients with APA or BHA who underwent AVS and dexamethasone-suppression NP-59 scintigraphy were included between 2010 and 2017. The correlation value between AVS and dexamethasone-suppression NP-59 scintigraphy was assessed using each lateralisation index (LIAVS and LI1NP-59). Tumour presence and size were evaluated using computed tomography. The sensitivity and specificity of dexamethasone-suppression NP-59 scintigraphy for APA according to each lateralisation index threshold were calculated. RESULTS: Of 29 patients, 12 presented with APA and 17 with BHA according to AVS. The correlation value between LIAVS and LI1NP-59 was 0.63 (P < 0.001). If the cut-off points were 2.55 and 1.80 in all cases, the sensitivity and specificity were 0.33 and 1.00 as well as 0.58 and 0.94, respectively. In adrenal microtumours (maximum diameter ≤10 mm), no cases revealed a cut-off point of >1.8. However, in adrenal macrotumours (maximum diameter >10 mm), the cut-off point of 2.55 represented the best compromise (sensitivity: 0.44; specificity: 1.00). CONCLUSION: NP-59 scintigraphy can be used as an alternative to AVS if there is a strong lateralisation on NP-59 scintigraphy and adrenal macrotumours observed on the computed tomography when AVS is technically challenging, particularly in the right adrenal vein cannulation, and if contraindications, such as allergy to contrast materials and renal failure, are observed.

Small intestinal bacterial overgrowth as a cause of protracted wound healing and vitamin D deficiency in a spinal cord injured patient with a sacral pressure sore: a case report.

BACKGROUND: Pressure sores are sometimes refractory to treatment, often due to malnutrition. Small intestinal bacterial overgrowth (SIBO) obstructs absorption in the digestive tract and causes malnutrition. However, little is known about the association between pressure sore wound healing and SIBO. Here, we report a case of a patient with a refractory sacral pressure sore and SIBO. CASE PRESENTATION: A 66-year-old woman who was spinal cord injured 14 years before visiting our hospital presented with the chief complaint of a sacral pressure sore, 10.0 × 6.5 cm in size, which was refractory to treatment. Physical examination showed abdominal distension and emaciation, with a body mass index of 15. Further examination revealed elevated serum alkaline phosphatase (1260 U/L), bilateral tibial fracture, multiple rib fracture, and osteoporosis. We diagnosed the patient with osteomalacia with vitamin D deficiency. Despite oral supplementation, serum levels of calcium, phosphorous, and vitamin D remained low. Also, despite concentrative wound therapy for the sacral pressure sore by plastic surgeons, no wound healing was achieved. Due to a suspicion of disturbances in nutrient absorption, we performed bacterial examination of collected gastric and duodenal fluid, which showed high numbers of bacteria in gastric content (104 E. coli, 105 Streptococcus species, and 105 Neisseria species) and duodenal content (106 E. coli, 104 Candida glabrata). Therefore, we diagnosed the patient with SIBO and started selective decontamination of the digestive tract using polymyxin B sulfate and amphotericin B. After starting treatment for SIBO, the sacral pressure sore began to heal and was nearly healed after 285 days. The patient's serum levels of calcium, phosphorous, vitamin D, and other fat-soluble vitamins also gradually increased after starting treatment for SIBO. CONCLUSION: We report a case of a patient with a refractory sacral pressure sore that healed after starting treatment for SIBO. We conclude that SIBO may be an overlooked cause of malnutrition and poor wound healing in patients with chronic pressure sores.

A Case of Hashimoto's Thyroiditis with Multiple Drug Resistance and High Expression of Efflux Transporters.

CONTEXT: Hashimoto's thyroiditis is the most common cause of hypothyroidism. Patients usually respond well to oral synthetic thyroxine (levothyroxine); however, for unknown reasons some individuals present with treatment-resistant Hashimoto thyroiditis. In cases of cancer and certain infectious diseases, the ATP binding cassette (ABC) transporters have been implicated in multidrug resistance, and we hypothesized and investigated a role of ABC transporters in drug-resistant Hashimoto's thyroiditis. CASE DESCRIPTION: The patient whose case we report had a history of Hashimoto's thyroiditis, immune thrombocytopenia, and refractory hypertension, with varying treatment resistance to the oral medications prescribed for each condition. In order to establish or exclude a genetic basis for her illness, we examined the patient's gene expression profiles using peripheral blood leukocytes, and found that ABCG2/BCRPexpression was significantly high compared with healthy volunteers. Also, the increased daunomycin efflux capacity of our patient's lymphocytes was successfully inhibited by fumitremorgin C, a specific ABCG2/BCRP inhibitor, and the patient's level of thyroid-stimulating hormone increased by 248.6% after administration of intact levothyroxine tablets but decreased by 45.1% when tablets were crushed. Her average blood pressure decreased from 166.3/108.5 mmHg to 125.9/78.8 mmHg when switching from intact to crushed losartan tablets. CONCLUSIONS: High expression and accelerated efflux transporter activity of ABCG2/BCRP in the small intestine are expected to contribute to the ineffectiveness of orally administered intact tablets in cases with treatment-resistant Hashimoto's thyroiditis, and crushed tablets can be more effective for some of these patients.

Aldosterone Reduction Rate After Saline Infusion Test May Be a Novel Prediction in Patients With Primary Aldosteronism.

OBJECTIVE: Accurate assessment and localization of aldosterone-producing adenomas (APAs) are essential for the treatment of primary aldosteronism (PA). Although adrenal venous sampling (AVS) is the standard method of reference for subtype diagnosis in PA, controversy exists concerning the criteria for its interpretation. This study aims to determine better indicators that can reliably predict subtypes of PA. METHOD: Retrospective, single-cohort analysis including 209 patients with PA who were subjected to AVS. Eighty-two patients whose plasma aldosterone concentrations (PAC) were normalized after surgery were histopathologically or genetically diagnosed with APA. The accuracy of image findings was compared to AVS results. Receiver operating characteristic (ROC) curve analysis between the operated and the no-apparent laterality groups was performed using AVS parameters and loading test for diagnosis of PA. RESULT: Agreement between image findings and AVS results was 56.3%. ROC curve analysis revealed that the lateralization index (LI) after adrenocorticotropin stimulation cutoff was 2.40, with 98.8% sensitivity and 97.1% specificity. The contralateral suppression index (CSI) cutoff value was 1.19, with 98.0% sensitivity and 93.9% specificity. All patients over the LI and CSI cutoff values exhibited unilateral subtypes. Among the loading test, the best classification accuracy was achieved using the PAC reduction rate after a saline infusion test (SIT) >33.8%, which yielded 87.2% sensitivity or a PAC after a SIT <87.9 pg/mL with 86.2% specificity for predicting bilateral PA. CONCLUSION: The combined criteria of the PAC reduction rate and PAC after the SIT can determine which subset of patients with APA who should be performed AVS for validation.

Adipose-Derived Stem Cells and Ceiling Culture-Derived Preadipocytes Cultured from Subcutaneous Fat Tissue Differ in Their Epigenetic Characteristics and Osteogenic Potential.

BACKGROUND: Adipose-derived stem cells and ceiling culture-derived preadipocytes can be harvested from subcutaneous adipose tissue. Little is known about the epigenetic differences, which may contribute to differences in osteogenic potential, between these cell types. The purpose of this study was to address the osteogenic potential and underlying epigenetic status of adipose-derived stem cells and ceiling culture-derived preadipocytes. METHODS: Adipose-derived stem cells and ceiling culture-derived preadipocytes were cultured from abdominal subcutaneous fat tissues of four metabolically healthy, lean female patients. After 7 weeks of culture, cellular responses to osteogenic differentiation media were examined. To evaluate the osteogenic potentials of undifferentiated adipose-derived stem cells and ceiling culture-derived preadipocytes, two types of epigenetic assessment were performed using next-generation sequencing: DNA methylation assays with the Human Methylation 450K BeadChip, and chromatin immunoprecipitation assays for trimethylation of histone H3 at lysine 4. RESULTS: Human ceiling culture-derived preadipocytes showed greater osteogenic differentiation ability than did adipose-derived stem cells. In an epigenetic survey of the promoters of four osteogenic regulator genes (RUNX2, SP7, ATF4, and BGLAP), the authors found a general trend toward decreased CpG methylation and increased trimethylation of histone H3 at lysine 4 levels in ceiling culture-derived preadipocytes as compared to adipose-derived stem cells, indicating that these genes were more likely to be highly expressed in ceiling culture-derived preadipocytes. CONCLUSIONS: The surveyed epigenetic differences between adipose-derived stem cells and ceiling culture-derived preadipocytes were consistent with the observed differences in osteogenic potential. These results enhance the authors' understanding of these cells and will facilitate their further application in regenerative medicine.

The role of tumor suppressor p53 in metabolism and energy regulation, and its implication in cancer and lifestyle-related diseases.

The tumor suppressor gene p53 is mutated in approximately more than 50% of human cancers. p53 is also referred to as the "cellular gatekeeper" or "guardian of the genome" because it protects the body from spreading mutated genome induced by various stress. When the cells receives stimuli such as DNA damage, oncogene activation, oxidative stress or undernutrition, p53 gives rise to a number of cellular responses, including cell cycle arrest, apoptosis, cellular senescence and metabolic adaptation. Related to energy metabolisms, it has been reported that p53 reduces glycolysis and enhances mitochondrial respiration. p53 is also involved in the regulation of other cellular metabolism and energy production systems: amino acid metabolism, fatty acid metabolism, nucleic acid metabolism, anti-oxidation, mitochondrial quality control, and autophagy. Moreover, recent studies have shown that p53 gene polymorphisms affect life expectancy and lifestyle-related disease such as type 2 diabetes, suggesting that there is a certain relationship between p53 function and metabolic disorders. In addition, mutant p53 protein does not only lose the tumor suppressor function, but it also gains novel oncogenic function and contributes to tumor development, involving cellular metabolism modification. Therefore, the importance of multifunctionality of p53, particularly with regard to intracellular metabolisms, arouses therapeutic interest and calls attention as the key molecule among cancer, lifestyle-related diseases and life expectancy.