3D Spheroids Facilitate Differentiation of Human Adipose-Derived Mesenchymal Stem Cells into Hepatocyte-Like Cells via p300-Mediated H3K56 Acetylation.

Hepatocyte-like cells (HLCs) that are differentiated from mesenchymal stem cells (MSCs) provide a valuable resource for drug screening and cell-based regeneration therapy. Differentiating HLCs into 3D spheroids enhances their phenotypes and functions. However, the molecular mechanisms underlying MSCs hepatogenic differentiation are not fully understood. In this study, we generated HLCs from human adipose-derived mesenchymal stem cells (hADMSCs) in both 2D and 3D cultures. We performed an acetyl-proteomics assay on the HLCs derived from both 2D and 3D differentiation and identified a differential change in H3K56 acetylation between the 2 differentiated cells. Our findings revealed that 3D differentiation activated ALB gene transcription by increasing the acetylation level of H3K56, thereby enhancing the phenotypes and functions of HLCs and further promoting their maturation. Notably, inhibiting p300 reduced the acetylation level of H3K56 during hepatogenic differentiation, leading to decreased phenotypes and functions of HLCs, whereas activation of p300 promoted hepatogenic differentiation, suggesting that p300 plays a critical role in this process. In summary, our study demonstrates a potential mechanism through which 3D spheroids differentiation facilitates hADMSCs differentiation into HLCs by promoting p300-mediated H3K56 acetylation, which could have significant clinical applications in liver regeneration and disease modeling.

Comparative Study of Two Common In Vitro Models for the Pancreatic Islet with MIN6.

BACKGROUND: Islet transplantation is currently considered the most promising method for treating insulin-dependent diabetes. The two most-studied artificial islets are alginate-encapsulated ß cells or ß cell spheroids. As three-dimensional (3D) models, both artificial islets have better insulin secretory functions and transplantation efficiencies than cells in two-dimensional (2D) monolayer culture. However, the effects of these two methods have not been compared yet. Therefore, in this study, cells from the mouse islet ß cell line Min6 were constructed as scaffold-free spheroids or alginate-encapsulated dispersed cells. METHODS: MIN6 cell spheroids were prepared by using Agarose-base microwell arrays. The insulin secretion level was determined by mouse insulin ELISA kit, and the gene and protein expression status of the MIN6 were performed by Quantitative polymerase chain reaction and immunoblot, respectively. RESULTS: Both 3D cultures effectively promoted the proliferation and glucose-stimulated insulin release (GSIS) of MIN6 cells compared to 2D adherent cells. Furthermore, 1% alginate-encapsulated MIN6 cells demonstrated more significant effects than the spheroids. In general, three pancreatic genes were expressed at higher levels in response to the 3D culture than to the 2D culture, and pancreatic/duodenal homeobox-1 (PDX1) expression was higher in the cells encapsulated in 1% alginate than that in the spheroids. A western blot analysis showed that 1% alginate-encapsulated MIN6 cells activated the phosphoinositide 3-kinase (PI3K)/serine/threonine protein kinase (AKT)/forkhead transcription factor FKHR (FoxO1) pathway more than the spheroids, 0.5% alginate-, or 2% alginate-encapsulated cells did. The 3D MIN6 culture, therefore, showed improved effects compared to the 2D culture, and the 1% alginate-encapsulated MIN6 cells exhibited better effects than the spheroids. The upregulation of PDX1 expression through the activation of the PI3K/AKT/FoxO1 pathway may mediate the improved cell proliferation and GSIS in 1% alginate-encapsulated MIN6 cells. CONCLUSION: This study may contribute to the construction of in vitro culture systems for pancreatic islets to meet clinical requirements.

Circulating Mucosal-Associated Invariant T Cells in a Large Cohort of Healthy Chinese Individuals From Newborn to Elderly.

Mucosal-associated invariant T (MAIT) cells, which are enriched in human blood and express a semi-invariant TCR chain, play important roles in conditions such as infectious diseases and cancer. The influence of age on levels and functional characteristics of circulating MAIT cells have not been fully addressed. Here we have collected blood samples from a large cohort of healthy Chinese individuals from newborn (cord blood) to the elderly and assessed the levels of circulating MAIT cells as well as their phenotype, activation and apoptosis status, and cytokine expression profiles after in vitro stimulation. We found that the frequencies of circulating MAIT cells gradually increased in blood from newborns as they progressed into adulthood (20-40 years old) but then decreased during further progression toward old age (>60 years old). The lowered numbers of circulating MAIT cells in the elderly was correlated with a gradual increase of apoptosis. A majority of circulating MAIT cells expressed the chemokine receptors CCR5 and CCR6, and most also expressed CD8 and CD45RO. Few expressed CD69 in cord blood, but the frequency increased with age. Upon in vitro activation with PMA plus ionomycin or IL12 plus IL18, fewer MAIT cells isolated from the young adult group expressed IFN-γ, IL17A and Granzyme B then cells from other age groups while the proportion of cells that expressed TNF-α was similar. Taken together, our data provide information for guiding the assessment of normal levels and phenotypes of MAIT cells at different ages in healthy individuals and patients.

A nanocomposite consisting of gold nanobipyramids and multiwalled carbon nanotubes for amperometric nonenzymatic sensing of glucose and hydrogen peroxide.

Gold nanobipyramids were synthesized by a seed-mediated growth method and then supported by multi-walled carbon nanotubes (denoted as AuNBP/MWCNTs). The electrocatalytic activity of the AuNBP/MWCNTs on a glassy carbon electrode (GCE) towards direct glucose oxidation and hydrogen peroxide reduction was superior to that of AuNBPs and MWCNTs. The modified GCE, operated at a typical working voltage of +0.15 V (vs. SCE) and in 0.1 M NaOH solution, exhibits a linear response in the 10 µM to 36.7 mM glucose concentration range with a 3.0 µM detection limit (at S/N = 3) and a sensitivity of 101.2 µA mM-1 cm-2. It also demonstrates good sensitivity towards hydrogen peroxide in at pH 7 solution at a working potential of -0.50 V (vs. SCE), with a linear response range from 5.0 µM to 47.3 mM, a sensitivity of 170.6 µA mM-1 cm-2 and a detection limit of 1.5 µM. Graphical abstract A electrochemical sensing platform based on the use of gold nanobipyramids and multi-walled carbon nanotubes nanocomposites (AuNBP/MWCNTs) is described for the determination of glucose and hydrogen peroxide.

Incidence and Distribution of Lobe-Specific Mediastinal Lymph Node Metastasis in Non-small Cell Lung Cancer: Data from 4511 Resected Cases.

OBJECTIVE: We aimed to investigate the incidence and distribution of mediastinal lymph node metastases (MLNM) in operable non-small cell lung cancer (NSCLC) with the purpose of guiding mediastinal lymph node dissection (MLND). METHODS: A total of 4511 NSCLC patients who underwent resection between January 2001 and December 2014 were included. These patients were preoperatively untreated and grouped according to the primary tumor lobes. The incidence and distribution of pathologic MLNM were compared among groups, and multivariate analysis was conducted to find the independent factors impacting MLNM. RESULTS: Lymph node involvement was observed in 1784 patients (39.5%). A total of 628 cases (13.9%) were N1-positive only, 752 cases (16.7%) were both N1- and N2-positive, and 404 cases (9.0%) were N2-positive only. The most common sites of mediastinal metastasis for different primary tumor lobes were the right upper lobe, station 4R (21.5%, 192/893); right middle lobe, station 7 (21.1%, 69/327); right lower lobe, station 7 (24.1%, 212/878); left upper lobe, station 5 (22.2%, 224/1008); and left lower lobe, station 7 (21.7%, 136/628). However, when only N2 cases were considered, each mediastinal lymph node zone can be involved with metastasis to a high proportion (> 5%). Multivariable analyses showed that poor cell differentiation, adenocarcinoma, larger tumor size, central type, and younger age were independent factors favoring MLNM. CONCLUSIONS: Different primary tumor locations have a different propensity to be sites of MLNM; however, once MLNM occurs, each zone can be involved and should not be neglected. Systematic MLND is the preferred procedure for operable NSCLC.

Efficacy of Endoscopic Ultrasonography for Determining Clinical T Category for Esophageal Squamous Cell Carcinoma: Data From 1434 Surgical Cases.

BACKGROUND: The efficacy of endoscopic ultrasonography (EUS) for determining T category is variable for esophageal squamous cell carcinoma (ESCC). We aimed to assess the efficacy of EUS in accurately identifying T category for ESCC based on the 8th AJCC Cancer Staging Manual. METHODS: A retrospective analysis was conducted using a prospectively collected ESCC database from January 2003 to December 2015, in which all patients underwent EUS examination followed by esophagectomy. The efficacy of EUS was evaluated by sensitivity, specificity, and accuracy compared with pathological T category as gold standard. Overall survival of different EUS-T (uT) categories was assessed. RESULTS: In total, 1434 patients were included, of whom 58.2% were correctly classified by EUS, with 17.9% being overstaged and 23.9% being understaged. The sensitivity and accuracy of EUS for Tis, T1a, T1b, T2, T3, and T4a categories were 15.8 and 98.8%, 16.3 and 95.7%, 33.1 and 89.3%, 56.8 and 65.0%, 65.8 and 70.0%, and 27.3 and 97.5%, respectively. The survival difference between uT1a and uT1b was not statistically significant (p = 0.90), nor was that between uT4a and uT4b (p = 0.34). However, when uT category was integrated as uTis, uT1, uT2, uT3, and uT4, overall survival was clearly distinguished between the categories (p < 0.01). CONCLUSIONS: EUS is in general feasible for classifying clinical T category for ESCC. However, EUS should be used with caution for discriminating between Tis, T1a, and T1b disease, as well as T4 disease.

Complete mitochondrial genome of a freshwater snail, Semisulcospira libertina (Cerithioidea: Semisulcospiridae).

The first complete mitochondrial genome (mitogenome) of Cerithioidea (Gastropoda: Caenogastropoda) was determined using a freshwater snail Semisulcospira libertina (Cerithioidea: Semisulcospiridae) as a representative species of the superfamily. The mitogenome was 15,432 bp in length, including 13 typical invertebrate protein-coding genes, 22 transfer RNA genes and 2 ribosomal RNA genes. The overall base composition was 31.4% for A, 17.8% for C, 34.8% for T and 16.0% for G with a A+T bias. The mitogenome of S. libertina displayed novel gene order arrangement compared with published Caenogastropoda mitogenomes to date. This mitogenome contributed in resolving phylogenetic position and interrelationships of Cerithioidea.

Gel-pad microarrays templated by patterned porous silicon for dual-mode detection of proteins.

A proof-of-concept study demonstrated the feasibility of a novel gel-pad microarray on porous silicon chips, by initiation of an atom transfer radical propagation (ATRP) polymerisation of (polyethylene glycol) methacrylate (PEGMA) with surface Si-H species, stepwise chemical conversions of the gel membrane to an NTA-Ni2+/histidine-tagged protein system, and matrix-assisted laser desorption/ionisation mass spectroscopy (MALDI MS) and fluorescence detections.