Generation of ventralized human thalamic organoids with thalamic reticular nucleus.

Human brain organoids provide unique platforms for modeling several aspects of human brain development and pathology. However, current brain organoid systems mostly lack the resolution to recapitulate the development of finer brain structures with subregional identity, including functionally distinct nuclei in the thalamus. Here, we report a method for converting human embryonic stem cells (hESCs) into ventral thalamic organoids (vThOs) with transcriptionally diverse nuclei identities. Notably, single-cell RNA sequencing revealed previously unachieved thalamic patterning with a thalamic reticular nucleus (TRN) signature, a GABAergic nucleus located in the ventral thalamus. Using vThOs, we explored the functions of TRN-specific, disease-associated genes patched domain containing 1 (PTCHD1) and receptor tyrosine-protein kinase (ERBB4) during human thalamic development. Perturbations in PTCHD1 or ERBB4 impaired neuronal functions in vThOs, albeit not affecting the overall thalamic lineage development. Together, vThOs present an experimental model for understanding nuclei-specific development and pathology in the thalamus of the human brain.

Epigallocatechin-3-gallate suppresses hemin-aggravated colon carcinogenesis through Nrf2-inhibited mitochondrial reactive oxygen species accumulation.

BACKGROUND: Previous studies have presented evidence to support the significant association between red meat intake and colon cancer, suggesting that heme iron plays a key role in colon carcinogenesis. Epigallocatechin-3-gallate (EGCG), the major constituent of green tea, exhibits anti-oxidative and anti-cancer effects. However, the effect of EGCG on red meat-associated colon carcinogenesis is not well understood. OBJECTIVES: We aimed to investigate the regulatory effects of hemin and EGCG on colon carcinogenesis and the underlying mechanism of action. METHODS: Hemin and EGCG were treated in Caco2 cells to perform the water-soluble tetrazolium salt-1 assay, lactate dehydrogenase release assay, reactive oxygen species (ROS) detection assay, real-time quantitative polymerase chain reaction and western blot. We investigated the regulatory effects of hemin and EGCG on an azoxymethane (AOM) and dextran sodium sulfate (DSS)-induced colon carcinogenesis mouse model. RESULTS: In Caco2 cells, hemin increased cell proliferation and the expression of cell cycle regulatory proteins, and ROS levels. EGCG suppressed hemin-induced cell proliferation and cell cycle regulatory protein expression as well as mitochondrial ROS accumulation. Hemin increased nuclear factor erythroid-2-related factor 2 (Nrf2) expression, but decreased Keap1 expression. EGCG enhanced hemin-induced Nrf2 and antioxidant gene expression. Nrf2 inhibitor reversed EGCG reduced cell proliferation and cell cycle regulatory protein expression. In AOM/DSS mice, hemin treatment induced hyperplastic changes in colon tissues, inhibited by EGCG supplementation. EGCG reduced the hemin-induced numbers of total aberrant crypts and malondialdehyde concentration in the AOM/DSS model. CONCLUSIONS: We demonstrated that EGCG reduced hemin-induced proliferation and colon carcinogenesis through Nrf2-inhibited mitochondrial ROS accumulation.

A pilot study on the effect of formulation and individual muscle mass on vitamin C absorption in randomized clinical study.

The current study investigated that the vitamin C absorption in plasma depends on the individual muscle mass and the formulation including drinks (Vita 500), capsules, and tablets by using a randomized and double-blind clinical study. The volunteers were divided into two groups that depended on their muscle mass, including those whose muscle mass was greater than 40% ( ≥ $ \ge $ 40%) and less than 40% muscle mass (<40%). Levels of vitamin C in blood plasma was analyzed by high-performance liquid chromatography by ultraviolet detection (HPLC-UV). The existing HPLC method was modified according to lab conditions but maintained a constantly low pH sample reduction procedure. The analytical method validated stability, linearity, recovery, reliability, and accuracy. The vitamin C absorption was the highest at 120 min after ingesting Vita 500 (21.47 ± 15.99 µmol/L). It was higher in the group that has more than 40% muscle mass compared to other formulations, such as tablets and capsules. The results from the current study indicate that vitamin C formulations differently affect the vitamin C absorption, and its effect depends on the muscle mass. As the results, liquid type vitamin C formulations could enhance vitamin C absorption, which resulted in an improvement of vitamin C absorption according to muscle mass. PRACTICAL APPLICATION: The results of this study may recommend using vitamin C supplementation as liquid type. It may also provide evidence that people with higher muscle mass can absorb vitamin C more efficiently.

Development of a patient-report pressure algometer for the quantification of abdominal examination.

BACKGROUND: Abdominal examination (AE), one of the primary diagnostic tools used in traditional Korean medicine (TKM), has a limitation of being subjective due to depending on individual practitioner's experience. Therefore, we devised a novel patient-report pressure algometer (PA) and performed a clinical trial to investigate its validity. METHODS: In total, 44 participants with functional dyspepsia and 44 healthy participants completed the study. The participants were allocated into one of two groups according to the existence of abdominal stiffness at 5 acupoints or abdominal tenderness at 12 acupoints diagnosed by TKM doctors. The pressure depth and pressure pain threshold (PPT) were evaluated using the PA at the same acupoints. We assessed the validity (sensitivity and specificity) of PA and calculated the area under the curve (AUC) and optimal cutoff value of the test variables (pressure depth and PPT) to criterion standards (abdominal stiffness and tenderness). RESULTS: Pressure depth and PPT assessed by PA showed high sensitivity and specificity in diagnosing abdominal stiffness and tenderness. The validity at CV-14 of diagnosing abdominal tenderness with PPT by PA had a sensitivity of 73.1%, specificity of 77.8%, and an AUC of 0.807 with a P value of < 0.001. CONCLUSION: This study may provide evidence of standardization and quantification of AE through PA.

PrPC Aptamer Conjugated-Gold Nanoparticles for Targeted Delivery of Doxorubicin to Colorectal Cancer Cells.

Anticancer drugs, such as fluorouracil (5-FU), oxaliplatin, and doxorubicin (Dox) are commonly used to treat colorectal cancer (CRC); however, owing to their low response rate and adverse effects, the development of efficient drug delivery systems (DDSs) is required. The cellular prion protein PrPC, which is a cell surface glycoprotein, has been demonstrated to be overexpressed in CRC, however, there has been no research on the development of PrPC-targeting DDSs for targeted drug delivery to CRC. In this study, PrPC aptamer (Apt)-conjugated gold nanoparticles (AuNPs) were synthesized for targeted delivery of Dox to CRC. Thiol-terminated PrPC-Apt was conjugated to AuNPs, followed by hybridization of its complementary DNA for drug loading. Finally, Dox was loaded onto the AuNPs to synthesize PrPC-Apt-functionalized doxorubicin-oligomer-AuNPs (PrPC-Apt DOA). The PrPC-Apt DOA were spherical nanoparticles with an average diameter of 20 nm. Treatment of CRC cells with PrPC-Apt DOA induced reactive oxygen species generation by decreasing catalase and superoxide dismutase activities. In addition, treatment with PrPC-Apt DOA inhibited mitochondrial functions by decreasing the expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha, complex 4 activity, and oxygen consumption rates. Compared to free Dox, PrPC-Apt DOA decreased proliferation and increased apoptosis of CRC cells to a greater degree. In this study, we demonstrated that PrPC-Apt DOA targeting could effectively deliver Dox to CRC cells. PrPC-Apt DOA can be used as a treatment for CRC, and have the potential to replace existing anticancer drugs, such as 5-FU, oxaliplatin, and Dox.

Steamed Ginger May Enhance Insulin Secretion through KATP Channel Closure in Pancreatic ß-Cells Potentially by Increasing 1-Dehydro-6-Gingerdione Content.

Ginger (Zingiber officinale Roscoe) and its active compounds (gingerols, shogaols and paradols) have been reported as having beneficial functions for several diseases, including diabetes. In this study, we revealed that the steaming process could enhance the anti-diabetic potential of ginger. To confirm the anti-diabetic effect of steamed ginger extract (GG03), we assessed pancreatic islets impaired by alloxan in zebrafish and demonstrated anti-hyperglycemic efficacy in a mouse model. The EC50 values of ginger extract (GE) and GG03 showed that the efficacy of GG03 was greater than that of GE. In addition, LC50 values demonstrated that GG03 had lower toxicity than GE, and the comparison of the Therapeutic Index (TI) proved that GG03 is a safer functional food. Furthermore, our data showed that GG03 significantly lowered hyperglycemia in a diabetic mouse model. HPLC was performed to confirm the change in the composition of steamed ginger. Interestingly, GG03 showed a 375% increase in 1-dehydro-6-gingerdione (GD) compared with GE. GD has not yet been studied much pharmacologically. Thus, we identified the protective effects of GD in the damaged pancreatic islets of diabetic zebrafish. We further assessed whether the anti-diabetic mechanism of action of GG03 and GD involves insulin secretion. Our results suggest that GG03 and GD might stimulate insulin secretion by the closure of KATP channels in pancreatic ß-cells.

hESC-Derived Thalamic Organoids Form Reciprocal Projections When Fused with Cortical Organoids.

Human brain organoid techniques have rapidly advanced to facilitate investigating human brain development and diseases. These efforts have largely focused on generating telencephalon due to its direct relevance in a variety of forebrain disorders. Despite its importance as a relay hub between cortex and peripheral tissues, the investigation of three-dimensional (3D) organoid models for the human thalamus has not been explored. Here, we describe a method to differentiate human embryonic stem cells (hESCs) to thalamic organoids (hThOs) that specifically recapitulate the development of thalamus. Single-cell RNA sequencing revealed a formation of distinct thalamic lineages, which diverge from telencephalic fate. Importantly, we developed a 3D system to create the reciprocal projections between thalamus and cortex by fusing the two distinct region-specific organoids representing the developing thalamus or cortex. Our study provides a platform for understanding human thalamic development and modeling circuit organizations and related disorders in the brain.

Effects of Lespedeza Cuneata aqueous extract on testosterone-induced prostatic hyperplasia.

CONTEXT: Lespedeza cuneata G. Don (Fabaceae), has been used as a traditional treatment of various diseases. There is a report L. cuneata effects on hormone replacement therapy for endocrine-related disease. However, studies related to benign prostatic hyperplasia (BPH) have not been investigated. OBJECTIVE: The effects of L. cuneata aqueous extract (LCW) on testosterone-induced prostatic hyperplasia (TPH) were examined. MATERIALS AND METHODS: Male Wistar rats (10 weeks, 330-350 g) were randomly divided to 6 groups (n = 6): Control group; TPH group (3 mg/kg, s.c, daily); TPH + LCW (25, 50, 100 mg/kg); TPH + Finasteride 10 mg/kg for 6 weeks. At the end of treatment, histological change of prostate, serum dihydrotestosterone (DHT) level, mRNA expression of 5α-reductase, inflammatory factors, proliferating cell nuclear antigen (PCNA) and fibroblast growth factor-2 (FGF-2) in prostate were examined. Then, LCW was treated with BPH-1, a human BPH cell line, at 25, 50, 100 µg/mL for 24 h and examine mRNA level of androgen receptor (AR) and prostate-specific antigen (PSA). In addition, the content of vicenin-2 was analyzed. RESULTS: LCW treatment of TPH inhibited serum DHT levels by 54.5, 51.2 and 54.1% and mRNA expression of 5α-reductase were inhibited 54.3, 61.3 and 73.6%, respectively. In addition, mRNA expression of inflammatory factors, PCNA and FGF-2 were decreased in the prostate of rats. Also, LCW attenuated mRNA level of AR and PSA in BPH-1 cell. The content of vicenin-2 in the LCW was analyzed to 0.89 mg/g. DISCUSSION AND CONCLUSIONS: Based on the results, LCW is a potential pharmacological candidate for the treatment of prostatic hyperplasia.

Protective effect of Lespedeza cuneata ethanol extract on Bisphenol A-induced testicular dysfunction in vivo and in vitro.

PURPOSE: Bisphenol A (BPA) has been regarded as a possible risk factor for reproductive health. We examined potential reproductive health benefits of Lespedeza cuneata ethanol extract (LCE). Previously, Lespedeza cuneata showed many therapeutic effects. However, the protective effect of LCE on BPA-induced testicular dysfunction and its mechanisms have not been precisely studied. METHODS: Mice were randomly divided into six groups (n = 7). Sperm counts and motility were measured by light microscope. Testosterone, total cholesterol, triglycerides, HDL, LDL-cholesterol, glucose, free fatty acids, hs-CRP, Angiotensinogen, Angiotensin II, GOT, GPT, TBARS, GSH, CAT, and SOD1 were measured in mouse serum. The potential protective effects of the LCE on mouse sertoli cells were evaluated. RESULTS: Oral administration of LCE in BPA-exposed male mice restored testis weight, sperm count, motility, and testosterone levels by inhibiting markers in serum. In addition, treatment with LCE in BPA-treated TM4 sertoli cells recovered cell viability by attenuating Bax expression and activating caspase 3 and PARP. CONCLUSIONS: These results indicate that LCE prevented BPA-induced testicular dysfunction and cell viability in BPA-treated TM4 sertoli cells. Our study also suggests that LCE has the potential to protect male reproductive health.

Distinct kinetics of inhibitory currents in thalamocortical neurons that arise from dendritic or axonal origin.

Thalamocortical neurons in the dorsal lateral geniculate nucleus (dLGN) transfer visual information from retina to primary visual cortex. This information is modulated by inhibitory input arising from local interneurons and thalamic reticular nucleus (TRN) neurons, leading to alterations of receptive field properties of thalamocortical neurons. Local GABAergic interneurons provide two distinct synaptic outputs: axonal (F1 terminals) and dendritic (F2 terminals) onto dLGN thalamocortical neurons. By contrast, TRN neurons provide only axonal output (F1 terminals) onto dLGN thalamocortical neurons. It is unclear if GABAA receptor-mediated currents originating from F1 and F2 terminals have different characteristics. In the present study, we examined multiple characteristics (rise time, slope, halfwidth and decay τ) of GABAA receptor-mediated miniature inhibitory postsynaptic synaptic currents (mIPSCs) originating from F1 and F2 terminals. The mIPSCs arising from F2 terminals showed slower kinetics relative to those from F1 terminals. Such differential kinetics of GABAAR-mediated responses could be an important role in temporal coding of visual signals.

Rivaroxaban vs Warfarin Sodium in the Ultra-Early Period After Atrial Fibrillation-Related Mild Ischemic Stroke: A Randomized Clinical Trial.

Importance: In atrial fibrillation (AF)-related acute ischemic stroke, the optimal oral anticoagulation strategy remains unclear. Objective: To test whether rivaroxaban or warfarin sodium is safer and more effective for preventing early recurrent stroke in patients with AF-related acute ischemic stroke. Design, Setting, and Participants: A randomized, multicenter, open-label, blinded end point evaluation, comparative phase 2 trial was conducted from April 28, 2014, to December 7, 2015, at 14 academic medical centers in South Korea among patients with mild AF-related stroke within the previous 5 days who were deemed suitable for early anticoagulation. Analysis was performed on a modified intent-to-treat basis. Interventions: Participants were randomized 1:1 to receive rivaroxaban, 10 mg/d for 5 days followed by 15 or 20 mg/d, or warfarin with a target international normalized ratio of 2.0-3.0, for 4 weeks. Main Outcomes and Measures: The primary end point was the composite of new ischemic lesion or new intracranial hemorrhage seen on results of magnetic resonance imaging at 4 weeks. Primary analysis was performed in patients who received at least 1 dose of study medications and completed follow-up magnetic resonance imaging. Key secondary end points were individual components of the primary end point and hospitalization length. Results: Of 195 patients randomized, 183 individuals (76 women and 107 men; mean [SD] age, 70.4 [10.4] years) completed magnetic resonance imaging follow-up and were included in the primary end point analysis. The rivaroxaban group (n = 95) and warfarin group (n = 88) showed no differences in the primary end point (47 [49.5%] vs 48 [54.5%]; relative risk, 0.91; 95% CI, 0.69-1.20; P = .49) or its individual components (new ischemic lesion: 28 [29.5%] vs 31 of 87 [35.6%]; relative risk, 0.83; 95% CI, 0.54-1.26; P = .38; new intracranial hemorrhage: 30 [31.6%] vs 25 of 87 [28.7%]; relative risk, 1.10; 95% CI, 0.70-1.71; P = .68). Each group had 1 clinical ischemic stroke, and all new intracranial hemorrhages were asymptomatic hemorrhagic transformations. Hospitalization length was reduced with rivaroxaban compared with warfarin (median, 4.0 days [interquartile range, 2.0-6.0 days] vs 6.0 days [interquartile range, 4.0-8.0]; P < .001). Conclusions and Relevance: In mild AF-related acute ischemic stroke, rivaroxaban and warfarin had comparable safety and efficacy. Trial Registration: clinicaltrials.gov Identifier: NCT02042534.

Hovenia Dulcis Extract Reduces Lipid Accumulation in Oleic Acid-Induced Steatosis of Hep G2 Cells via Activation of AMPK and PPARα/CPT-1 Pathway and in Acute Hyperlipidemia Mouse Model.

Hovenia dulcis Thunb. (HDT) was known to have anti-fatigue, anti-diabetes, neuroprotective, and hepatoprotective effects. In the present study, the anti-fatty liver mechanism of HDT was elucidated in oleic acid (OA)-treated Hep G2 cells and acute hyperlipidemia mouse model using Triton WR-1339. Here, HDT activated p-AMP-activated protein kinase (p-AMPK), proliferator activated receptor-α, carnitine palmitoyltransferase and also inhibited the expression of lipogenesis and cholesterol synthesis proteins, such as 3-hydroxy-3-methylglutaryl-CoA reductase, sterol regulatory element binding protein-1c, SREBP-2, and fatty acid synthase in OA-treated Hep G2 cells. Conversely, AMPK inhibitor compound C blocked the anti-fatty liver effect of HDT to induce AMPK phosphorylation and decrease 3-hydroxy-3-methylglutaryl-CoA reductase and lipid accumulation by oil red O staining in OA-treated Hep G2 cells. Additionally, HDT pretreatment protected against the increase of serum total cholesterol, triglyceride, low-density lipoprotein cholesterol and phospholipid in an acute hyperlipidemia mouse model with enhancement of glutathione reductase, glutathione peroxidase, superoxide dismutase, and catalase activities. Taken together, HDT inhibits OA-induced hepatic lipid accumulation via activation of AMPK and proliferator activated receptor-α/carnitine palmitoyltransferase signaling and enhancement of antioxidant activity as a potent candidate for nonalcoholic fatty liver disease and hyperlipidemia. Copyright © 2016 John Wiley & Sons, Ltd.

Antioxidant effects of Cirsium setidens extract on oxidative stress in human mesenchymal stem cells.

Human mesenchymal stem cells (MSCs) may be used in cell-based therapy to promote neovascularization for the treatment of ischemic diseases. However, high levels of reactive oxygen species (ROS) derived from the pathophysiological ischemic environment induce senescence and apoptosis of MSCs, resulting in reduced functionality and defective neovascularization. Therefore, the present study aimed to determine the protective effects of Cirsium setidens, a natural product, on oxidative stress­induced apoptosis in MSCs. The present study investigated for the change of ROS levels in MSCs using ROS assays. In addition, cell viability determined by MTT and TUNEL assays. Western blot analysis was performed to investigate the change of apoptosis­associated proteins in MSCs. Treatment of MSCs with hydrogen peroxide (H2O2; 200 µM) significantly increased intracellular ROS levels and cell death; however, pretreatment with C. setidens (100 µg/ml) suppressed H2O2­induced ROS generation and increased the survival of MSCs. H2O2­induced ROS production increased the levels of phosphorylated­p38 mitogen activated protein kinase, c­Jun N­terminal kinase, ataxia telangiectasia mutated and p53; these increases were inhibited by pretreatment with C. setidens. In addition, C. setidens inhibited ROS­induced apoptosis of MSCs by increasing the expression levels of the anti­apoptotic protein B­cell lymphoma 2 (BCL­2), and decreasing the expression levels of the proapoptotic protein BCL­2­associated X protein. These findings indicated that pretreatment of MSCs with C. setidens may prevent ROS­induced oxidative injury by regulating the oxidative stress­associated signaling pathway, and suppressing the apoptosis­associated signal pathway. Therefore, C. setidens may be developed as a beneficial broad­spectrum agent for enhancing the effectiveness of MSC transplantation in the treatment of ischemic diseases.

Hematopoietic effect of fermented deer antler extract in iron deficient diet-induced anemic rats.

OBJECTIVE: To assess the hematopoietic effects of fermented deer antler extract using a dietinduced anemic animal model to facilitate the utilization of fermented deer antler extract and its derived products. METHODS: Thirty 3-week-old female Sprague-Dawley rats were treated for 5 weeks. The rats were randomly divided into 6 groups and treated as follows: control, saline; NFA200, non-fermented deer antler extract 200 mg/kg; NFA500, non-fermented deer antler extract 500 mg/kg; FAB200, fermented deer antler extract 200 mg/kg; FAB500, fermented deer antler extract 500 mg/kg; and PC, heme iron 0.2 mg/kg. Blood parameters, iron content in the liver and spleen, hepatic δ-aminolevulinic acid dehydrogenase (ALAD) activity and divalent metal transporter 1 (DMT1) mRNA expression were analyzed. RESULTS: No detectable significant differences were observed in blood parameters among groups. The decrease in the hepatic ALAD activity in anemic rats was significantly improved by fermented deer antler extract supplementation (P<0.05); however, non-fermented deer antler extract supplementation did not result in a significant improvement (P>0.05). The hepatic DMT1 mRNA expression level was increased significantly by supplementation with both the fermented deer antler extract and the non-fermented deer antler extract in a dose-dependent manner compared with nontreatment in anemic rats (P<0.05). CONCLUSION: The hematopoietic activity induced by deer antler extract in dietinduced anemic rats might be increased through the fermentation process.

Doxycycline supplementation allows for the culture of human ESCs/iPSCs with media changes at 3-day intervals.

Culturing human embryonic stem and induced pluripotent stem cells (hESCs/iPSCs) is one of the most costly and labor-intensive tissue cultures, as media containing expensive factors/cytokines should be changed every day to maintain and propagate undifferentiated hESCs/iPSCs in vitro. We recently reported that doxycycline, an anti-bacterial agent, had dramatic effects on hESC/iPSC survival and promoted self-renewal. In this study, we extended the effects of doxycycline to a more practical issue to save cost and labor in hESC/iPSC cultures. Regardless of cultured cell conditions, hESCs/iPSCs in doxycycline-supplemented media were viable and proliferating for at least 3 days without media change, while none or few viable cells were detected in the absence of doxycycline in the same conditions. Thus, hESCs/iPSCs supplemented with doxycycline can be cultured for a long period of time with media changes at 3-day intervals without altering their self-renewal and pluripotent properties, indicating that doxycycline supplementation can reduce the frequency of media changes and the amount of media required by 1/3. These findings strongly encourage the use of doxycycline to save cost and labor in culturing hESCs/iPSCs.

Fucoidan protects mesenchymal stem cells against oxidative stress and enhances vascular regeneration in a murine hindlimb ischemia model.

BACKGROUND: Mesenchymal stem cells (MSCs) have the potential to differentiate into multiple cell lineages. Given this potential for tissue regeneration, MSC-based therapeutic applications have been considered in recent years. However, ischemia-induced apoptosis has been reported to be one of the main causes of MSC death following transplantation. The primary objective of this study was to determine whether a natural antioxidant, fucoidan, could protect MSCs from ischemia-induced apoptosis in vitro and in vivo. Furthermore, we investigated the mechanism of action of fucoidan's anti-ischemic effect in MSCs. METHODS AND RESULT: Pre-treatment with fucoidan (10 µg/mL) suppressed the increase in H2O2-induced reactive oxygen species (ROS) levels and drastically reduced apoptotic cell death in MSCs. Fucoidan inhibited the activation of the pro-apoptotic proteins p38-mitogen-activated protein kinase (MAPK), Jun N-terminal kinase (JNK), and caspase-3, and augmented the expression of the anti-apoptosis protein cellular inhibitor of apoptosis (cIAP). Moreover, fucoidan significantly increased manganese superoxide dismutase (MnSOD) expression and decreased cellular ROS levels via the Akt pathway, resulting in enhanced cell survival. In a murine hindlimb ischemia model, transplanted fucoidan-treated MSCs showed significantly enhanced cell survival and proliferation in ischemic tissues. Functional recovery and limb salvage also remarkably improved in mice injected with fucoidan-stimulated MSCs compared with mice injected with non-stimulated MSCs. CONCLUSION: Taken together, these results show that fucoidan protects MSCs from ischemia-induced cell death by modulation of apoptosis-associated proteins and cellular ROS levels through regulation of the MnSOD and Akt pathways, suggesting that fucoidan could be powerful therapeutic adjuvant for MSC-based therapy in ischemic diseases.

Vitamin C facilitates dopamine neuron differentiation in fetal midbrain through TET1- and JMJD3-dependent epigenetic control manner.

Intracellular Vitamin C (VC) is maintained at high levels in the developing brain by the activity of sodium-dependent VC transporter 2 (Svct2), suggesting specific VC functions in brain development. A role of VC as a cofactor for Fe(II)-2-oxoglutarate-dependent dioxygenases has recently been suggested. We show that VC supplementation in neural stem cell cultures derived from embryonic midbrains greatly enhanced differentiation toward midbrain-type dopamine (mDA) neurons, the neuronal subtype associated with Parkinson's disease. VC induced gain of 5-hydroxymethylcytosine (5hmC) and loss of H3K27m3 in DA phenotype gene promoters, which are catalyzed by Tet1 and Jmjd3, respectively. Consequently, VC enhanced DA phenotype gene transcriptions in the progenitors by Nurr1, a transcription factor critical for mDA neuron development, to be more accessible to the gene promoters. Further mechanism studies including Tet1 and Jmjd3 knockdown/inhibition experiments revealed that both the 5hmC and H3K27m3 changes, specifically in the progenitor cells, are indispensible for the VC-mediated mDA neuron differentiation. We finally show that in Svct2 knockout mouse embryos, mDA neuron formation in the developing midbrain decreased along with the 5hmC/H3k27m3 changes. These findings together indicate an epigenetic role of VC in midbrain DA neuron development.

Doxycycline enhances survival and self-renewal of human pluripotent stem cells.

We here report that doxycycline, an antibacterial agent, exerts dramatic effects on human embryonic stem and induced pluripotent stem cells (hESC/iPSCs) survival and self-renewal. The survival-promoting effect was also manifest in cultures of neural stem cells (NSCs) derived from hESC/iPSCs. These doxycycline effects are not associated with its antibacterial action, but mediated by direct activation of a PI3K-AKT intracellular signal. These findings indicate doxycycline as a useful supplement for stem cell cultures, facilitating their growth and maintenance.

Toward integrated molecular diagnostic system (i MDx): principles and applications.

Integrated molecular diagnostic systems ( iMDx), which are automated, sensitive, specific, user-friendly, robust, rapid, easy-to-use, and portable, can revolutionize future medicine. This review will first focus on the components of sample extraction, preservation, and filtration necessary for all point-of-care devices to include for practical use. Subsequently, we will look for low-powered and precise methods for both sample amplification and signal transduction, going in-depth to the details behind their principles. The final field of total device integration and its application to the clinical field will also be addressed to discuss the practicality for future patient care. We envision that microfluidic systems hold the potential to breakthrough the number of problems brought into the field of medical diagnosis today.

Cell-based high-throughput odorant screening system through visualization on a microwell array.

The development of a cell-based high-throughput screening system has attracted much attention from researchers who study drug screening mechanisms and characterization of G-protein coupled receptors (GPCRs). Although olfactory receptors (ORs) constitute the largest group of GPCRs that play a critical role recognizing and discriminating odorants, only a few ORs have been characterized, and most remain orphan. The conventional cell-based assay system for characterizing GPCRs, including ORs, is very laborious, time consuming, and requires an expensive assay system. In this study, we developed a simple, low-cost miniaturized odorant screening method by combining Micro-Electro-Mechanical system (MEMs) technique and visualization technique for detecting an odorant response. We fabricated PEG microwell from a photocrosslinkable polyethylene glycol diacrylate (PEGDA) solution and applied it to cell culture and a reverse transfection platform for cell-based high-throughput screening. For the first time, the olfactory receptors were expressed on the microwell platform using reverse transfection technique. The various olfactory receptors can be expressed simultaneously using this technique and the microwell spotted with olfactory receptor genes can be used as a high-throughput screening platform. The odorant response was detected via fluorescence analysis on the microwell using a cAMP response element (CRE) reporter assay. We tested this platform using four de-orphaned ORs. This new cell-based screening method not only reduced numerous time-consuming steps but also allowed for simple, efficient, and quantitative screening and patterning of large numbers of GPCRs including ORs, which can help to visualize the OR response to odorants on a microwell.