Other SectionsABSTRACTINTRODUCTIONCONCEPT OF ORGANOID-BASED REGENERATIVE MEDICINETISSUE SPECIFIC ORGANOID-BASED REGENERATIVE MEDICINECONCLUSIONACKNOWLEDGEMENTSCONFLICTS OF INTERESTFIGURESTABLEREFERENCES
Background: There are no reliable biomarkers to accurately differentiate indolent thyroid tumors from more aggressive thyroid cancers. This study aimed to develop new DNA methylation markers for diagnosis and recurrence risk stratification of papillary thyroid carcinoma (PTC). Methods: Thyroid tumor-specific DNA methylation profiling was investigated in 34 fresh frozen tissues, which included nontumor (n = 7), noninvasive follicular thyroid neoplasms with papillary-like nuclear features (NIFTP, n = 6) and PTC (n = 21), using the Illumina HumanMethylation EPIC array. We performed a genome-wide assessment of thyroid tumor-specific differentially methylated CpG sites in the discovery set, then validated the top candidate markers in an independent set of 293 paraffin tissue samples comprised of follicular adenoma (FA, n = 61), Hürthle cell adenoma (HA, n = 24), NIFTP (n = 56), PTC (n = 120), follicular thyroid carcinoma (n = 27), and Hürthle cell carcinoma (n = 5), by pyrosequencing. Results: Three selected markers (cg10705422, cg17707274, and cg26849382) differentiated nonmalignant (FA, HA, and NIFTP) tumors from differentiated thyroid cancers with area under the receiver operating characteristic curve of 0.83, 0.83, and 0.80, respectively. Low DNA methylation levels for three markers were significantly associated with recurrent or persistent disease (odds ratio (OR) = 3.860 [95% confidence interval (CI) 1.194-12.475]) and distant metastasis (OR = 4.009 [CI 1.098-14.632]) in patients with differentiated thyroid cancer. A subgroup analysis for the validation set showed that PTC patients with low DNA methylation levels more frequently had aggressive histology, extrathyroidal extension, lymph node metastasis, BRAFV600E mutations, and recurrent or persistent disease than those with high levels of methylation markers. All PTC patients who developed disease recurrence had low DNA methylation levels for three markers. Conclusions: DNA methylation levels of three markers can be useful for differentiating differentiated thyroid cancer from nonmalignant follicular thyroid lesions, and may serve as prognostic biomarkers for predicting recurrent or persistent disease after surgery for differentiated thyroid cancer.
Pathological changes in the epigenetic landscape of chromatin are hallmarks of cancer. Our previous study showed that global methylation of promoters may increase or decrease during the transition from gastric mucosa to intestinal metaplasia (IM) to gastric cancer (GC). Here, CpG hypomethylation of the serine/threonine kinase STK31 promoter in IM and GC was detected in a reduced representation bisulfite sequencing database. STK31 hypomethylation, which resulted in its upregulation in 120 cases of primary GC, was confirmed. Using public genome‑wide histone modification data, upregulation of STK31 promoter activity was detected in primary GC but not in normal mucosae, suggesting that STK31 may be repressed in gastric mucosa but activated in GC as a consequence of hypomethylation‑associated chromatin remodeling. STK31 knockdown suppressed the proliferation, colony formation and migration activities of GC cells in vitro, whereas stable overexpression of STK31 promoted the proliferation, colony formation, and migration activities of GC cells in vitro and tumorigenesis in nude mice. Patients with GC in which STK31 was upregulated exhibited significantly shorter survival times in a combined cohort. Thus, activation of STK31 by chromatin remodeling may be associated with gastric carcinogenesis and also may help predict GC prognosis.
Pathological changes in the epigenetic landscape of chromatin are hallmarks of cancer. The caudal-type homeobox gene CDX2 is not expressed in normal gastric epithelia but rather in adult intestinal epithelia, and it is overexpressed in intestinal metaplasia (IM). However, it remains unclear how CDX2 transcription is suppressed in normal gastric epithelial cells and overexpressed in IM. Here, we demonstrate that methylation of the CDX2 promoter increases with age in Helicobacter pylori-positive, noncancerous gastric tissue, whereas the promoter is demethylated in paired gastric tumors in which CDX2 is upregulated. Moreover, we also found that the CDX2 promoter is demethylated in IM as well as gastric tumor. Immunohistochemistry revealed that CDX2 is present in foci of parts of the gastric mucosae but highly expressed in IM as well as in gastric tumors, suggesting that the elevated level of CDX2 in IM and gastric tumors may be attributable to promoter demethylation. Our data suggest that CDX2 repression may be associated with promoter methylation in noncancerous H. pylori-positive mucosa but its upregulation might be attributable to increased promoter activity mediated by chromatin remodeling during gastric carcinogenesis.
Colistin (polymyxin E) is an antibiotic that is effective against multidrug-resistant gram-negative bacteria. However, the high incidence of nephrotoxicity caused by colistin limits its clinical use. To identify compounds that might ameliorate colistin-induced nephrotoxicity, we obtained 1707 compounds from the Korea Chemical Bank and used a high-content screening (HCS) imaging-based assay. In this way, we found that bimatoprost (one of prostaglandin F2α analogue) ameliorated colistin-induced nephrotoxicity. To further assess the effects of bimatoprost on colistin-induced nephrotoxicity, we used in vitro and in vivo models. In cultured human proximal tubular cells (HK-2), colistin induced dose-dependent cytotoxicity. The number of terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL)-positive cells, indicative of apoptosis, was higher in colistin-treated cells, but this effect of colistin was ameliorated by cotreatment with bimatoprost. The generation of reactive oxygen species, assessed using 2,7-dichlorodihydrofluorescein diacetate, was less marked in cells treated with both colistin and bimatoprost than in those treated with colistin alone. Female C57BL/6 mice (n = 10 per group) that were intraperitoneally injected with colistin (10 mg/kg/12 hr) for 14 days showed high blood urea nitrogen and serum creatinine concentrations that were reduced by the coadministration of bimatoprost (0.5 mg/kg/12 hr). In addition, kidney injury molecule-1 (KIM1) and Neutrophil gelatinase-associated lipocalin (NGAL) expression also reduced by bimatoprost administration. Further investigation in tubuloid and kidney organoids also showed that bimatoprost attenuated the nephrotoxicity by colistin, showing dose-dependent reducing effect of KIM1 expression. In this study, we have identified bimatoprost, prostaglandin F2α analogue as a drug that ameliorates colistin-induced nephrotoxicity.
This study investigates the impact of sodium channel protein type 1 subunit alpha (SCN1A) gene knockout (SCN1A KO) on brain development and function using cerebral organoids coupled with a multiomics approach. From comprehensive omics analyses, we found that SCN1A KO organoids exhibit decreased growth, dysregulated neurotransmitter levels, and altered lipidomic, proteomic, and transcriptomic profiles compared to controls under matrix-free differentiation conditions. Neurochemical analysis reveals reduced levels of key neurotransmitters, and lipidomic analysis highlights changes in ether phospholipids and sphingomyelin. Furthermore, quantitative profiling of the SCN1A KO organoid proteome shows perturbations in cholesterol metabolism and sodium ion transportation, potentially affecting synaptic transmission. These findings suggest dysregulation of cholesterol metabolism and sodium ion transport, with implications for synaptic transmission. Overall, these insights shed light on the molecular mechanisms underlying SCN1A-associated disorders, such as Dravet syndrome, and offer potential avenues for therapeutic intervention.
Abstract Background Xerostomia is a pathological condition characterized by decreased salivation due to salivary gland dysfunction and is frequently attributed to irreversible damage as a side effect of radiation therapy. Stem cell–derived organoid therapy has garnered attention as a promising avenue for resolving this issue. However, Matrigel, a hydrogel commonly used in organoid culture, is considered inappropriate for clinical use due to its undefined composition and immunogenicity. In this study, we aimed to develop a method for culturing collagen-based human salivary gland organoids (hSGOs) suitable for clinical applications and evaluated their therapeutic effectiveness. Methods Human salivary gland stem cells were isolated from the salivary gland tissues and cultured in both Matrigel and collagen. We compared the gene and protein expression patterns of salivary gland–specific markers and measured amylase activity in the two types of hSGOs. To evaluate the therapeutic effects, we performed xenogeneic and allogeneic transplantation using human and mouse salivary gland organoids (hSGOs and mSGOs), respectively, in a mouse model of radiation-induced xerostomia. Results hSGOs cultured in Matrigel exhibited self-renewal capacity and differentiated into acinar and ductal cell lineages. In collagen, they maintained a comparable self-renewal ability and more closely replicated the characteristics of salivary gland tissue following differentiation. Upon xenotransplantation of collagen-based hSGOs, we observed engraftment, which was verified by detecting human-specific nucleoli and E-cadherin expression. The expression of mucins, especially MUC5B, within the transplanted hSGOs suggested a potential improvement in the salivary composition. Moreover, the allograft procedure using mSGOs led to increased salivation, validating the efficacy of our approach. Conclusions This study showed that collagen-based hSGOs can be used appropriately in clinical settings and demonstrated the effectiveness of an allograft procedure. Our research has laid the groundwork for the future application of collagen-based hSGOs in allogeneic clinical trials.
The study was carried out to examine the anti-obesity effects of 40% ethanol extract from green tea seed (GS) and quantitative determination of caffeine as its major compound. The specificity was satisfied with retention time and UV spectrum by analysis of caffeine using HPLC and comparison with standard compound. It showed a high linearity in the calibration curve with a coefficient of correlation (R²) of 0.9974. The amount of caffeine in GS was about 4.649 mg/g (0.465%) in the three times analysis, and relative standard deviation (RSD) was less than 0.452% by the validated method. The anti-obesity effects of GS were evaluated by using Oil Red O staining in 3T3-L1 adipocytes and body weight, visceral fat and lipid profiles in high fat diet (HFD)-induced C57BL/6 obese mice. Our results indicated that treatment with GS dose-dependently decreased lipid accumulation contents (p0.001). Moreover, after oral administration for 30 days feeding with HFD-induced obses mice, GS (100 and 300 mg/kg/day) produced a significant decrease in serum total cholesterol (TC), glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT) and visceral fat. Thus, the result of this study indicate that the GS may be a useful resource for the management of obesity.
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