Rabbit Meat Extract Induces Browning in 3T3-L1 Adipocytes via the AMP-Activated Protein Kinase Pathway.

The browning of white adipocytes may be an innovative approach to address obesity. This study investigated the effects of rabbit meat extract on 3T3-L1 adipocytes, with a specific emphasis on inducing browning. The browning effects of rabbit meat extract were evaluated by analyzing genes specifically expressed in 3T3-L1 adipocytes using quantitative PCR and immunoblotting. Rabbit meat extract increased the expression of brown adipocyte-specific markers, UCP1 and PGC1α, and mitochondrial biogenesis factors, TFAM and NRF1, without affecting cell viability in fully differentiated 3T3-L1 adipocytes. Moreover, adipocyte differentiation and the triglyceride content were decreased; hormone-sensitive lipase activity was promoted. Rabbit meat extract activated the AMPK pathway in the differentiated 3T3-L1 cells. However, in adipocytes treated with rabbit meat extract, the expression of genes related to browning was reduced by the AMP-activated protein kinase (AMPK) inhibitor, dorsomorphin dihydrochloride. To the best of our knowledge, this is the first study to demonstrate that rabbit meat extract induces the browning of white adipocytes via the activation of the AMPK pathway, thereby demonstrating its therapeutic potential in preventing obesity.

Effect of Different Gas-Stunning Conditions on Heme Pigment Solutions and on the Color of Blood, Meat, and Small Intestine of Rabbits.

The objectives of this study were to elucidate the effect of different high-concentration levels of inert gases (Ar and N2) on heme-pigment solutions and on the color of the blood, meat, and small intestine of rabbits; and to figure out the proper concentration level of inert gas (Ar or N2) for the complete stunning of the rabbit. To observe the changing of color attributes, a research study was conducted in the abattoir of the National Institute of Animal Science (NIAS), RDA, Republic of Korea. This experiment had 2 parts, (i) A trial on heme-pigment solutions (hemoglobin and myoglobin solution) was conducted in a gas chamber with different high-concentration levels of carbon dioxide, argon, nitrogen, and normal air; and (ii) a rabbit experiment was conducted­stunning with different high-concentration levels of carbon dioxide, argon, nitrogen, and the Halal method (non-stunning). A small-size digital gas chamber was used for this experiment (size: width 850 mm × depth 1350 mm). Artificial hemoglobin (Hb) and myoglobin (Mb) solutions were created from "porcine hemoglobin lyophilized powder" and "equine skeletal muscle myoglobin lyophilized powder", respectively. In the heme-pigment solutions trial, 10 treatments were used: (i) 80% carbon dioxide (T1), (ii) 85% carbon dioxide (T2), (iii) 90% carbon dioxide (T3), (iv) 80% argon (T4), (v) 85% argon (T5), (vi) 90% argon (T6), (vii) 80% nitogen (T7), (viii) 85% nitogen (T8), (ix) 90% nitogen (T9), and (x) normal air (T10). Heme-pigment solutions (both Hb and Mb) were exposed with each treatment for four separate durations of time (30 s, 1 min, 2 min, and 4 min); and every sample (Hb and Mb) was exposed during each duration of time for 10 times (n = 10). In the rabbit experiment, seven treatments were used (i) 85% carbon dioxide (T1), (ii) 90% carbon dioxide (T2), (iii) 85% argon (T3), (iv) 90% argon (T4), (v) 96% nitrogen (T5), (vi) 98% nitogen (T6), and (vii) the Halal method (non-stunning) (T7). Forty-two rabbits (mixed-breed) were collected from the nearest commercial farm and randomly selected for a treatment group (n = 6). The average body weight was 2.3 kg. For stunning, each individual rabbit was separately kept in a gas chamber, after which each specific gas was used to fulfill the desired level in the pit. After reaching the desired level of concentration, rabbits were stunned within a very short time. It was observed that the time required for stunning in the T1, T2, T3, T4, T5 and T6 treatment was 79−82, 68−73, 97−103, 88−91, 164−173, and 108−115 s, respectively. In the case of the Halal method (non-stunning), as per the rules of Islam, rabbits were slaughtered without stunning. After slaughtering, in all treatments, the blood, meat, and small intestine of each animal were collected carefully and kept in a cool room in which the temperature was −2 °C, and after 24 h, the color attributes­lightness (L*), redness (a*), and yellowness (b*)­were measured via a Chroma Meter. It was found that in both experiments (trial on heme-pigment solutions and rabbit stunning), the color values (L*, a* and b*) showed a significant difference (p < 0.05) among the treatment groups. The CO2-treated groups showed high redness (a*) and low lightness (L*) and yellowness (b*), which indicated a dark-red color, and N2-treated groups showed high lightness (L*) and yellowness (b*) and low redness (a*), which implied a bright-red color. The effect of the Ar was in between the CO2- and N2-treated groups. These phenomena were observed both in the heme-pigment solutions (Hb and Mb) and in the blood, meat, and small intestine of the rabbits. N2-stunned animals fulfill the fundamental desire of consumers to purchase bright red-colored fresh meat. Therefore, in view of color attributes, consumer satisfaction, and animal welfare, N2 gas can be thought of as a valuable alternative to stunning. Considering the time required for complete stunning and desirable color attributes, a 98% concentration of N2 is best for rabbit stunning. As such, it could be used as a better option for the gas stunning of animals.

Effects of High Concentration Nitrogen Gas Stunning of Pigs on the Quality Traits of Meat and Small Intestine.

The objective of present study was to investigate the feasibility of utilizing only high concentration of nitrogen gas in the stunning of pigs and its effects on the quality traits of the meat and small intestine.To conduct this experiment, three treatment groups were compared: (i) electric stunning (T1), (ii) CO2 (80%) gas stunning (T2), and (iii) N2 (98%) gas stunning (T3). A total of 21 standard pigs (Landrace × Yorkshire × Duroc; LYD) were collected from a commercial pig farm, randomly selecting seven pigs for each group (body weight of 104.5 to 120.6 kg). For stunning, each individual pig was separately kept in a gas chamber, after which each specific gas was used to fulfill the desired level in the pit. To obtain the desired level of concentration for each gas (N2 at 98% and CO2 at 80%), approximately 80 min and 35 min were required, respectively. It was observed that after reaching the desired level of concentration, pigs were stunned within a very short time (for CO2, 90 s and for N2, 120 s). For electric stunning, standard quality electric devices were used. After slaughtering, the meat and small intestine of each animal was collected separately and kept in a cool room where temperature was −2 °C. In the meat and small intestine, L* (Lightness) and b* (Yellowness) were high (p < 0.05) in the T1 and T3 groups. The T2 group showed high a* (Redness) (p < 0.05) values in both the meat and small intestine. A proximate composition of meat showed no significant differences except moisture. The water holding capacity (WHC), cooking loss (CL), and Warner-Bratzler shear force (WBSF) of meat were lowest in the T2 group, but not at a notable difference compared to T3. In the small intestine, L* (Lightness), a* (Redness), b* (Yellowness), and thickness significantly differed (p < 0.05) in each group, but WBSF showed no significance between the T2 and T3 groups. It is concluded that a high concentration of N2 gas (98%) may be considered in the stunning of pigs, and its effect on meat and small intestine is favorable.

Quality Properties of Dry-Aged Beef (Hanwoo Cattle) Crust on Pork Patties.

This study evaluated the effects of crust derived from dry-aged beef (Hanwoo cattle) on the quality of pork patties. Pork patty samples were prepared with different amounts of crust (0­control, 1, 2, and 3%). The protein, fat, and ash contents in the crust samples were significantly higher than those in the control sample (p < 0.05). The CIE b* value of uncooked pork patties with crust added was significantly lower than that of the control patties (p < 0.05). The pH and CIE L* values of uncooked patty batter samples decreased with increasing concentrations of crust (p < 0.05). However, the viscosity increased proportionally with an increase in crust (p < 0.05). Samples containing 3% crust showed significantly higher uncooked and cooked CIE a*, water-holding capacity, cooking yield, and shear force than the control sample (p < 0.05). Moreover, samples containing 2% and 3% crust showed significantly lower diameter and thickness reductions than those of the control sample (p < 0.05). The sensory evaluation conferred by the crust was significantly higher than that of the control sample (p < 0.05). Overall, our results suggest that pork patties supplemented with 3% crust have improved properties.

Effect of Hanwoo Crust on the Physicochemical Properties of Emulsion-Type Sausages.

This study aimed to investigate the effects of Hanwoo crust, inedible surface layer formed during dry aging, on the physicochemical properties of emulsion-type sausages. Sausage samples were prepared with various amounts of Hanwoo crust-0% (i.e., control), 1%, 2%, and 3%. The physicochemical properties studied included the proximate composition, pH, color, water holding capacity (WHC), cooking yield (CY), and viscosity. Texture profile analysis (TPA) and sensory evaluation were also carried out. Protein, fat, and ash contents of the Hanwoo crust-treated samples were found to be significantly higher than those of the control (p<0.05). Moreover, the CIE b* value of cooked sausage with Hanwoo crust treatments was significantly lower than that of the control (p<0.05). The CIE L* value of uncooked and cooked samples with 3% Hanwoo crust was significantly lower than that of the control (p<0.05). In contrast, the CIE a* value of uncooked and cooked samples with 3% Hanwoo crust was significantly higher than that of the control (p<0.05). The viscosity of the uncooked samples increased with increasing Hanwoo crust content. Samples containing 3% Hanwoo crust exhibited significantly higher WHC and CY than the control (p<0.05). In the TPA, samples containing 2% and 3% Hanwoo crust showed significantly higher hardness, gumminess, and chewiness than the control (p<0.05). Overall, the sensory properties of Hanwoo crust treatments were significantly better than those of the control (p<0.05). In conclusion, adding 3% Hanwoo crust to emulsion-type sausage leads to optimal physicochemical properties.

Olig2 regulates p53-mediated apoptosis, migration and invasion of melanoma cells.

Melanoma is a disease with a high recurrence rate and poor prognosis; therefore, the need for targeted therapeutics is steadily increasing. Oligodendrocyte transcription factor2 (Olig2) is a basic helix-loop-helix transcription factor that is expressed in the central nervous system during embryonic development. Olig2 is overexpressed in various malignant cell lines such as lung carcinoma, glioma and melanoma. Olig2 is known as a key transcription factor that promotes tumor growth in malignant glioma. However, the role of Olig2 in melanoma is not well characterized. We analyzed the role of Olig2 in apoptosis, migration, and invasion of melanoma cells. We confirmed that Olig2 was overexpressed in melanoma cells and tissues. Reduction of Olig2 increased apoptosis in melanoma cells by increasing p53 level and caspase-3/-7 enzyme activity. In addition, downregulation of Olig2 suppressed migration and invasion of melanoma cells by inhibiting EMT. Reduction of Olig2 inhibited expression of MMP-1 and the enzyme activity of MMP-2/-9 induced by TGF-ß. Moreover, Olig2 was involved in the downstream stages of MEK/ERK and PI3K/AKT, which are major signaling pathways in metastatic progression of melanoma. In conclusion, this study demonstrated the crucial roles of Olig2 in apoptosis, migration, and invasion of melanoma and may help to further our understanding of the relationship between Olig2 and melanoma progression.

Evaluation of interradicular space, soft tissue, and hard tissue of the posterior palatal alveolar process for orthodontic mini-implant, using cone-beam computed tomography.

INTRODUCTION: To investigate the anatomy of the posterior palatal alveolar process, which is often used for placement of the orthodontic mini-implant (OMI), and to suggest simple guidelines for safe placement of OMI. METHODS: Cone-beam computed tomography (CBCT) scans of 60 patients (30 men, 30 women; age range, 18-39 years; average age, 25.8 years) was used to measure the palatal interradicular distance, the palatal bone thickness, and the palatal soft-tissue thickness. Measurements were performed on the area from the maxillary canine to the maxillary second molar based on the vertical distance apical from the cementoenamel junction. The CBCT data were analyzed by Bonferroni correction for multiple testing and the multivariable mixed linear model. RESULTS: The palatal interradicular distance was the widest between the second premolar and the first molar and the narrowest between the first and second premolars. The palatal bone thickness at interdental sites was the thickest between the first and second premolars and the thinnest between the first and second molars. The interdental palatal soft-tissue thickness from the canine to the second premolar was thicker than any other area. There were minor measurement differences between genders and positive correlations between vertical distance from the cementoenamel junction plane and all of the parameters. CONCLUSION: In this study, we evaluated the anatomy of the posterior palatal area using CBCT scans of adult patients. The data will provide guidelines to the clinicians before OMI placement in the posterior palatal alveolar process.

Physicochemical properties of crust derived from dry-aged Holstein and Hanwoo loin.

This study evaluated the quality characteristics of crust derived from dry-aged Holstein and Hanwoo loins and their effects on food as additives. With respect to physicochemical properties, we examined the proximate composition, pH value, salinity, color, water and fat absorption, emulsifying capacity, and swelling yield. The protein and ash contents in the Holstein crust were significantly higher than those in the Hanwoo crust (p < 0.0001). The fat content in the Hanwoo crust was significantly higher than that in the Holstein crust (p < 0.01). The salinity, lightness, and yellowness of the Hanwoo crust were significantly lower than those of the Holstein crust (p < 0.001). Furthermore, the pH value and emulsifying capacity of the Hanwoo crust were significantly higher than those of the Holstein crust (p < 0.001). The fat absorption of the Holstein crust was significantly higher than that of the Hanwoo crust (p < 0.001). The swelling yield of the Holstein crust was significantly higher than that of the Hanwoo crust at pH 3 and 4 (p < 0.001), whereas the swelling yield of the Hanwoo crust was significantly higher than that of the Holstein crust at pH 7 (p < 0.001). Principal component analysis of dry-aged Hanwoo, Holstein, and non-aged Holstein showed different flavor patterns for each sample. Finally, the results showed that the crusts derived from dry-aged Hanwoo and Holstein loins were suitable flavor enhancers.

16-Kauren-2-beta-18,19-triol inhibits melanosome transport in melanocytes by down-regulation of melanophilin expression.

BACKGROUND: Rab27a, Mlph, and MyoVa form a tripartite complex and relate to melanosome distribution. Melanophilin (Mlph) acts as a linker protein between Rab27a and MyoVa. The biological activity and function of 16-kauren on the expression of Mlph has not yet been studied. OBJECTIVE: We examined the effect of 16-kauren on melanosome transport and skin pigmentation. METHODS: Murine Melan-a melanocytes and SP-1 keratinocytes were used for in vitro analysis. Western blot analysis, quantitative real-time polymerase chain reaction, luciferase assay and immunohistochemical staining in 3D pigmented human skin model were performed. RESULTS: We found that 16-kauren inhibits melanosome transport in Melan-a melanocytes without affecting melanin synthesis. Treatment with 16-kauren reduced melanophilin (Mlph), a key protein in melanosome transport, in Melan-a melanocytes, at both the protein and mRNA levels while it did not affect the expression of Rab27a and MyoVa, the other two key proteins for melanosome transport. Notably, the expression of melanogenic proteins, including tyrosinase, trp1, trp2, and MITF, was not affected by 16-kauren. However, 16-kauren attenuated melanosome distribution in co-culture of Melan-a melanocytes and SP-1 keratinocytes as well as in Melan-a monolayer culture. In further confirmation of the depigmenting effects of 16-kauren on Melanoderm™, a 3D pigmented human skin model, treatment with 16-kauren for 12 days increased the brightness of the tissue as determined by lightness value and reduced the distribution of melanosomes as shown in histological examination. CONCLUSION: These results demonstrated that 16-kauren is a selective modulator of a melangenic target, Mlph expression, and can be employed as a new depigmenting strategy.

Identification of MicroRNA Targeting Mlph and Affecting Melanosome Transport.

Melanosomes undergo a complex maturation process and migrate into keratinocytes. Melanophilin (Mlph), a protein complex involving myosin Va (MyoVa) and Rab27a, enables the movement of melanosomes in melanocytes. In this study, we found six miRNAs targeting Mlph in mouse using two programs (http://targetscan.org and DianaTools). When melan-a melanocytes were treated with six synthesized microRNAs, miR-342-5p, miR-1839-5p, and miR-3082-5p inhibited melanosome transport and induced melanosome aggregation around the nucleus. The other microRNAs, miR-5110, miR-3090-3p, and miR-186-5p, did not inhibit melanosome transport. Further, miR-342-5p, miR-1839-5p, and miR-3082-5p decreased Mlph expression. The effect of miR-342-5p was the strongest among the six synthesized miRNAs. It inhibited melanosome transport in melan-a melanocytes and reduced Mlph expression in mRNA and protein levels in a dose-dependent manner; however, it did not affect Rab27a and MyoVa expressions, which are associated with melanosome transport. To examine miR-342-5p specificity, we performed luciferase assays in a mouse melanocyte-transfected reporter vector including Mlph at the 3'-UTR (untranslated region). When treated with miR-342-5p, luciferase activity that had been reduced by approximately 50% was restored after inhibitor treatment. Therefore, we identified a novel miRNA affecting Mlph and melanosome transport, and these results can be used for understanding Mlph expression and skin pigmentation regulation.

Autotransplantation of premolars and space closure in a patient with inflamed sinuses.

Congenital absence of permanent teeth can be treated by means of dental implants and prosthetics. Tooth autotransplantation is an alternative in growing patients because continued eruption of the transplanted tooth and associated alveolar bone growth can be expected. This clinical report presents tooth autotransplantation in a 10-year-old boy with chronic maxillary rhinosinusitis, diagnosed by the department of otorhinolaryngology. The patient's mandibular second premolars were transplanted to a congenitally edentulous maxillary premolar region. There was insufficient alveolar bone during transplantation because of pneumatization, but normal root development with vertical alveolar bone growth was observed during a 3-year follow-up. Healing of the transplant in the right side without closing of the apex and without signs of obliteration after 4 years is exceptional.

Fabrication and characterization of periodically patterned silica fiber structures for enhanced second-order nonlinearity: publisher's note.

This publisher's note amends the author list of [Opt. Express23, 8113 (2015)].

Postoperative complications of microvascular decompression for hemifacial spasm: lessons from experience of 2040 cases.

Microvascular decompression (MVD) is an effective and safe treatment option that offers the prospect of definitive cure for hemifacial spasm (HFS). However, there are potential risks of complications for MVD associated with retromastoid suboccipital craniectomy (RmSOC) and cranial nerves in particular. The purpose of this study was to identify clinical characteristics of possible complications after MVD for HFS and to establish appropriate management concept of these complications. We retrospectively reviewed medical records of 2040 patients who underwent RmSOC with MVD for HFS at Samsung medical center between January 1998 and March 2013. Of 2040 patients, 2027 were followed (99.4%). Of the 2027 patients, 1841 (90.8%) exhibited complete relief or minimal symptoms, and 113 (5.6%) reported improved spasm but had mild remnant symptoms. After operation, the most frequently developed complications were facial nerve palsy (8.19%), followed by middle ear effusion (4.90%) and hearing loss (3.63%). There were two cases of supratentorial subdural hemorrhage, three cases of infarction. MVD was found to be safe and effective treatment for HFS, in consistent with previous reports. Some of the complications such as facial nerve palsy, middle ear effusion, and hearing loss are relatively common. However, they have mild clinical courses that are usually transient.

A new possible mechanism of hearing loss after microvascular decompression for hemifacial spasm.

OBJECTIVE: Hearing loss is a well-known complication that may occur during microvascular decompression (MVD) for hemifacial spasm (HFS). Cause and risk factors are highly variable. We present cases of hearing loss induced by saline overinfusion after MVD. STUDY DESIGN: Retrospective review in a tertiary referral center. INTERVENTION: Three hundred thirty-one patients with HFS underwent MVD from March 2009 to October 2010. MAIN OUTCOME MEASURES: Brain stem auditory evoked potential (BAEP) was monitored during the surgery. Before completion of the dural closure, the surgical field was routinely filled with warm saline to avoid postoperative pneumocephalus and epidural hematoma. RESULTS: Seven patients experienced a change in wave V amplitude and latency after the dural closure. In 2 patients, the amplitudes decreased by less than 50%, and latencies were delayed by less than 1.0 ms, ipsilaterally in 1 patient and contralaterally in the other. In 1 patient, decreased amplitude and delayed latency appeared bilaterally with more severity on the operated side, accompanied by delayed ipsilateral permanent hearing loss. In 4 of the 7 patients, an ipsilateral response of BAEP was completely absent. Of these 4 patients, 2 experienced permanent hearing loss, and another 2 patients who underwent dural reopening and saline drainage had restoration of their normal hearing. CONCLUSION: Intradural compression due to overinfusion of saline may lead to postoperative hearing loss, although the incidence is low, and immediate decompression by drainage may be required.

Dual-energy CT to detect recurrent HCC after TACE: initial experience of color-coded iodine CT imaging.

OBJECTIVES: To evaluate the feasibility of diagnosing recurrence of HCC after TACE color-coded iodine CT (CICT) based on arterial phase scans obtained by a dual-energy CT (DECT) scanner. METHODS: A CICT scan was acquired from an iodine map after applying material decomposition of the liver tissue and setting a threshold attenuation level for viable tumors. Two radiologists reviewed both conventional and CICT sets in 31 patients who had a history of TACE for HCC. The performances in detecting local tumor progression (LTP) were evaluated by alternative free-response receiver operating characteristics. The rate of uncertain diagnosis and interobserver agreement of the diagnosis were explored. Additionally, the reading time and radiation dose were also investigated. RESULTS: The mean figures of merit of the conventional and CICT sets for LTP were 0.818 and 0.847, respectively (p=0.459). The rate of uncertain diagnosis was significantly decreased in CICT sets (34.5% vs. 0%), and interobserver agreement was improved (k=0.527 vs. 0.718). On the CICT set, mean reading time was reduced by 49s and mean radiation dose was also decreased by 18.3% when replacing the non-contrast CT with CICT. CONCLUSIONS: CICT is comparable to conventional liver CT protocol in demonstrating viable HCCs, while it allows a reduction in radiation dose.

Delayed cranial nerve palsy after microvascular decompression for hemifacial spasm.

OBJECTIVE: Microvascular decompression (MVD) for hemifacial spasm (HFS) is a safe and effective treatment with favorable outcomes. The purpose of this study was to evaluate the incidence of delayed cranirve ( VI, VII, and VIII ) palsy following MVD and its clinical courses. METHODS: Between January 1998 and December 2009, 1354 patients underwent MVD for HFS at our institution. Of them, 100 patients (7.4%) experienced delayed facial palsy (DFP), one developed sixth nerve palsy, and one patient had delayed hearing loss. RESULTS: DFP occurred between postoperative day number 2 and 23 (average 11 days). Ninety-two patients (92%) completely recovered; however, House-Brackmann grade II facial weakness remained in eight other patients (8%). The time to recovery averaged 64 days (range, 16 days to 9 months). Delayed isolated sixth nerve palsy recovered spontaneously without any medical or surgical treatment after 8 weeks, while delayed hearing loss did not improve. CONCLUSION: Delayed cranial nerve (VI, VII, and VIII) palsies can occur following uncomplicated MVD for HFS. DFP is not an unusual complication after MVD, and prognosis is fairly good. Delayed sixth nerve palsy and delayed hearing loss are extremely rare complications after MVD for HFS. We should consider the possibility of development of these complications during the follow up for MVD.

Effects of yoga exercise on serum adiponectin and metabolic syndrome factors in obese postmenopausal women.

OBJECTIVE: Regular and continuous yoga exercise is one of the most important nonpharmacological methods of improving serum lipid concentrations, adipose tissue, and metabolic syndrome factors. The purpose of this study was to analyze the effects of yoga exercise on serum adiponectin and metabolic syndrome factors in obese postmenopausal Korean women. METHODS: Sixteen healthy postmenopausal women aged 54.50 ± 2.75 years with more than 36% body fat were randomly assigned to either a yoga exercise group (n = 8) or to a "no exercise" control group (n = 8). The variables of body composition, visceral fat, serum adiponectin, and metabolic syndrome factors were measured in all the participants before and after the 16-week study. RESULTS: Body weight, percentage of body fat, lean body mass, body mass index, waist circumference, and visceral fat area had significantly decreased. High-density lipoprotein cholesterol and adiponectin had significantly increased, but total cholesterol, triglyceride, low-density lipoprotein cholesterol, blood pressure, insulin, glucose, and homoeostasis model assessment-insulin resistance had significantly decreased. Serum adiponectin concentrations were significantly correlated with waist circumference, high-density lipoprotein cholesterol, diastolic blood pressure, and homoeostasis model assessment-insulin resistance in the postyoga exercise group. CONCLUSIONS: Our findings indicate that yoga exercise improves adiponectin level, serum lipids, and metabolic syndrome risk factors in obese postmenopausal women. Consequently, yoga exercise will be effective in preventing cardiovascular disease caused by obesity in obese postmenopausal Korean women.

Disease-specific induced pluripotent stem cells: a platform for human disease modeling and drug discovery.

The generation of disease-specific induced pluripotent stem cell (iPSC) lines from patients with incurable diseases is a promising approach for studying disease mechanisms and drug screening. Such innovation enables to obtain autologous cell sources in regenerative medicine. Herein, we report the generation and characterization of iPSCs from fibroblasts of patients with sporadic or familial diseases, including Parkinson's disease (PD), Alzheimer's disease (AD), juvenile-onset, type I diabetes mellitus (JDM), and Duchenne type muscular dystrophy (DMD), as well as from normal human fibroblasts (WT). As an example to modeling disease using disease-specific iPSCs, we also discuss the previously established childhood cerebral adrenoleukodystrophy (CCALD)- and adrenomyeloneuropathy (AMN)-iPSCs by our group. Through DNA fingerprinting analysis, the origins of generated disease-specific iPSC lines were identified. Each iPSC line exhibited an intense alkaline phosphatase activity, expression of pluripotent markers, and the potential to differentiate into all three embryonic germ layers: the ectoderm, endoderm, and mesoderm. Expression of endogenous pluripotent markers and downregulation of retrovirus-delivered transgenes [OCT4 (POU5F1), SOX2, KLF4, and c-MYC] were observed in the generated iPSCs. Collectively, our results demonstrated that disease-specific iPSC lines characteristically resembled hESC lines. Furthermore, we were able to differentiate PD-iPSCs, one of the disease-specific-iPSC lines we generated, into dopaminergic (DA) neurons, the cell type mostly affected by PD. These PD-specific DA neurons along with other examples of cell models derived from disease-specific iPSCs would provide a powerful platform for examining the pathophysiology of relevant diseases at the cellular and molecular levels and for developing new drugs and therapeutic regimens.

Induced pluripotent stem cell models from X-linked adrenoleukodystrophy patients.

OBJECTIVE: Because of a lack of an appropriate animal model system and the inaccessibility of human oligodendrocytes in vivo, X-linked adrenoleukodystrophy (X-ALD)-induced pluripotent stem cells (iPSCs) would provide a unique cellular model for studying etiopathophysiology and development of therapeutics for X-ALD. METHODS: We generated and characterized iPSCs of the 2 major types of X-ALD, childhood cerebral ALD (CCALD) and adrenomyeloneuropathy (AMN), and differentiated them into oligodendrocytes and neurons. We evaluated disease-relevant phenotypes by pharmacological and genetic approaches. RESULTS: We established iPSCs from the patients with CCALD and AMN. Both CCALD and AMN iPSCs normally differentiated into oligodendrocytes, the cell type primarily affected in the X-ALD brain, indicating no developmental defect due to the ABCD1 mutations. Although low in X-ALD iPSCs, very long chain fatty acid (VLCFA) level was significantly increased after oligodendrocyte differentiation. VLCFA accumulation was much higher in CCALD oligodendrocytes than AMN oligodendrocytes but was not significantly different between CCALD and AMN neurons, indicating that the severe clinical manifestations in CCALD might be associated with abnormal VLCFA accumulation in oligodendrocytes. Furthermore, the abnormal accumulation of VLCFA in the X-ALD oligodendrocytes can be reduced by the upregulated ABCD2 gene expression after treatment with lovastatin or 4-phenylbutyrate. INTERPRETATION: X-ALD iPSC model recapitulates the key events of disease development (ie, VLCFA accumulation in oligodendrocytes), provides new clues for better understanding of the disease, and allows for early and accurate diagnosis of the disease subtypes. X-ALD oligodendrocytes can be a useful cell model system to develop new therapeutics for treating X-ALD.