Tzyy-Wen Chiou

Single dose intravenous thioacetamide administration as a model of acute liver damage in rats

Thioacetamide (TAA) has been used extensively in the development of animal models of acute liver injury. Frequently, TAA is administered intraperitoneally to induce liver damage under anaesthesia. However, it is rarely administered by intravenous injection in conscious rats. The experiments in this study were designed to induce acute liver damage by single intravenous injection of TAA (0, 70 and 280 mg/kg) in unrestrained rats. Biochemical parameters and cytokines measured during the 60‐h period following TAA administration, included white blood cells (WBC), haemoglobulin (Hb), platelet, aspartate transferase (GOT), alanine transferase (GPT), total bilirubin (TBIL), direct bilirubin (DBI), albumin, ammonia (NH3), r‐glutamyl transpeptidase (r‐GT), tumour necrosis factor‐α (TNF‐α) and interleukin‐6 (IL‐6). Rats were sacrificed by decapitation 60 h after TAA administration and livers were removed immediately for pathology and immunohistochemical (IHC) examination. Another group of rats were sacrificed by decapitation 1, 6 and 24 h after TAA administration and livers were removed immediately for time course change of pathology and IHC examination. TAA significantly increased blood WBC, GOT, GPT, TBIL, DBIL, NH3, r‐GT, TNF‐α and IL‐6 levels but decreased the blood Hb, platelet and albumin level. The levels of histopathological damage in the liver after intravenous TAA administration were also increased with a dose‐dependent trend and more increased at 60 h after TAA administration. The levels of inducible nitric oxide synthase (iNOS) and nuclear factor‐κB (NF‐κB) detected by IHC in the liver after intravenous TAA administration were also increased with a dose‐dependent trend and more increased at 1 h after TAA administration. Single intravenous TAA administration without anaesthesia is a restorable animal model which may be used to investigate acute liver damage.

Overexpression of the orphan receptor Nur77 and its translocation induced by PCH4 may inhibit malignant glioma cell growth and induce cell apoptosis.

In previous study, n‐butylidenephthalide (BP), a natural compound from Angelica sinensis, has anti‐glioblastoma multiform (GBM) cell effects. In this study, we modified BP structure to increase anti‐GBM cell effects. The anti‐GBM cell effects of one derivative of BP, (Z)‐N‐(2‐(dimethylamino)ethyl)‐2‐(3‐((3‐oxoisobenzofuran‐1(3H)‐ylidene)methyl)phenoxy)acetamide (PCH4) were tested in vitro and in vivo.

Inhibitory effect of n-butylidenephthalide on neointimal hyperplasia in balloon injured rat carotid artery.

This investigation was designed to determine the inhibitory effects and mechanisms of n‐butylidenephthalide (BP) from Angelica sinensis on smooth muscle cell (SMC) proliferation in vitro and in balloon injured rat carotid artery. Treatment of cultured rat aorta SMC‐derived A7r5 cells with 25–100 μg/mL BP significantly inhibited the proliferation and arrested the cell cycle in G0/G1 phase. BP induced the expression and migration of Nur77 from the nucleus to the cytoplasm. Among signal pathways, JNK and p38 MAPK were phosphorylated after BP treatment. In vivo, the neointimal area of common carotid artery 2 weeks after balloon injury reduced significantly in Sprague‐Dawley rats treated with 150–300 mg/kg BP compared with the control. The proliferative activity indicated by immunohistochemical detection of Ki‐67 positive cells in the neointima was significantly decreased in the 60–300 mg/kg BP treatment groups. The apoptotic activity indicated by cleaved caspase‐3 positive cells and Nur77 positive cells in the neointima was significantly increased in rats treated with 60–300 mg/kg BP. This study demonstrated BP inhibited neointimal hyperplasia in balloon injured rat carotid artery due to its dual effects of proliferative inhibition and apoptotic induction on SMCs. Up‐regulation of Nur77 gene may partly explain the antihyperplasia activity of BP on the neointima. Copyright

n-Butylidenephthalide exhibits protection against neurotoxicity through regulation of tryptophan 2, 3 dioxygenase in spinocerebellar ataxia type 3

&NA; Spinocerebellar ataxia type 3 or Machado‐Joseph disease (SCA3/MJD) is characterized by the repetition of a CAG codon in the ataxin‐3 gene (ATXN3), which leads to the formation of an elongated mutant ATXN3 protein that can neither be denatured nor undergo proteolysis in the normal manner. This abnormal proteolysis leads to the accumulation of cleaved fragments, which have been identified as toxic and further they act as a seed for more aggregate formation, thereby increasing toxicity in neuronal cells. To date, there have been few studies or treatment strategies that have focused on controlling toxic fragment formation. The aim of this study is to develop a potential treatment strategy for addressing the complications of toxic fragment formation and to provide an alternative treatment strategy for SCA3. Our preliminary data on anti‐aggregation and toxic fragment formation using an HEK (human embryonic kidney cells) 293T‐84Q‐eGFP (green fluorescent protein) cell model identified n‐butylidenephthalide (n‐BP) as a potential drug treatment for SCA3. n‐BP decreased toxic fragment formation in both SCA3 cell and animal models. Moreover, results showed that n‐BP can improve gait, motor coordination, and activity in SCA3 mice. To comprehend the molecular basis behind the control of toxic fragment formation, we used microarray analysis to identify tryptophan metabolism as a major player in controlling the fate of mutant ATXN3 aggregates. We also demonstrated that n‐BP functions by regulating the early part of the kynurenine pathway through the downregulation of tryptophan 2, 3‐dioxygenase (TDO2), which decreases the downstream neurotoxic product, quinolinic acid (QA). In addition, through the control of TDO2, n‐BP also decreases active calpain levels, an important enzyme involved in the proteolysis of mutant ATXN3, thereby decreasing toxic fragment formation and associated neurotoxicity. Collectively, these findings indicate a correlation between n‐BP, TDO2, QA, calpain, and toxic fragment formation. Thus, this study contributes to a better understanding of the molecular interactions involved in SCA3, and provides a novel potential treatment strategy for this neurodegenerative disease. Graphical abstract Figure. No caption available. Highlightsn‐BP has anti‐aggregating and pro‐proliferative effect which aid in improvement of the pathological state of SCA3 mice.In SCA3, TDO2 has been observed to be upregulated which in turn upregulates downstream product QA.QA being an agonist of NMDA‐R further may increase the influx of Ca2+ ions which increases Ca 2+ dependent calpain activity.Calpain through proteolysis cleaves mutant ATXN3 to form toxic fragment.Hence, n‐BP may decrease toxic fragment formation through its activity on TDO2 and improve the SCA3 neuropathology.

Antitumor and Antimicrobial Activity of Some Cyclic Tetrapeptides and Tripeptides Derived from Marine Bacteria

Marine derived cyclo(Gly-l-Ser-l-Pro-l-Glu) was selected as a lead to evaluate antitumor-antibiotic activity. Histidine was chosen to replace the serine residue to form cyclo(Gly-l-His-l-Pro-l-Glu). Cyclic tetrapeptides (CtetPs) were then synthesized using a solution phase method, and subjected to antitumor and antibiotic assays. The benzyl group protected CtetPs derivatives, showed better activity against antibiotic-resistant Staphylococcus aureus in the range of 60–120 μM. Benzyl group protected CtetPs 3 and 4, exhibited antitumor activity against several cell lines at a concentration of 80–108 μM. However, shortening the size of the ring to the cyclic tripeptide (CtriP) scaffold, cyclo(Gly-l-Ser-l-Pro), cyclo(Ser-l-Pro-l-Glu) and their analogues showed no antibiotic or antitumor activity. This phenomenon can be explained from their backbone structures.

The antisenescence effect of Trans-cinnamaldehyde on adipose-derived stem cells

As assuring cell quality is an essential parameter for the success of stem cell therapy, the impact of various senescence-inducing stress signals, and strategies to circumvent them, has been an important area of focus in stem cell research. The aim of this study was to demonstrate the capacity of trans-cinnamaldehyde (TC) in reversing stress-induced senescence and maintaining the quality of stem cells in a chemically (H2O2)-induced cell senescence model. Because of the availability and the promising application potential in regenerative medicine, adipose-derived stem cells (ADSCs) were chosen for the study. We found that H2O2 treatment resulted in the expression of senescence characteristics in the ADSCs, including decreased proliferation rate, increased senescence-associated β-galactosidase (SA-β-gal) activity, decreased silent mating type information regulation 2 homolog (SIRT1) expression, and decreased telomerase activity. However, TC treatment was sufficient to rescue or reduce the effects of H2O2 induction, ultimately leading to an increased proliferation rate, a decrease in the percentage of SA-β-gal-positive cells, upregulation of SIRT1 expression, and increased telomerase activity of the senescent ADSCs at the cellular level. Moreover, a chemically induced liver fibrosis animal model was used to evaluate the functionality of these rescued cells in vivo. Liver dysfunction was established by injecting 200 mg/kg thioacetamide (TAA) intraperitoneally into Wistar rats every third day for 60 days. The experimental rats were separated into groups: normal group (rats without TAA induction), sham group (without ADSC transplantation), positive control group (transplanted with normal ADSCs), H2O2 group (transplanted with H2O2-induced senescent ADSCs), and H2O2 + TC group (transplanted with ADSCs pretreated with H2O2 and then further treated with TC). In the transplantation group, 1 × 106 human ADSCs were introduced into each rat via direct liver injection. Based on the biochemical analysis and immunohistochemical staining results, it was determined that the therapeutic effects on liver fibrosis by the induced senescent ADSCs (H2O2 group) were not as significant as those exerted by the normal ADSCs (the positive control group). However, the H2O2 + TC group showed significant reversal of liver damage when compared to the H2O2 group 1 week posttransplantation. These data confirmed that the TC treatment had the potential to reduce the effects of H2O2-induced senescence and to restore in vivo functionality of the induced senescent ADSCs. It is therefore suggested that TC has potential applications in maintaining the quality of stem cells and could aid in treating senescence-related disorders.

An Evidence-based Perspective of Angelica Sinensis (Chinese Angelica) for Cancer Patients

The main compounds in Angelica sinensis (Chinese angelica) acetone extract AS-C are ferulic acid, ligustilide, brefeldin A, butylidenephthalide, as well as polysaccharides. Polysaccharide have been determined their effects on various human cancer cells. Subsequently, the active component of AS-C, butylidenephthalide (BP), has been investigated for its antitumor effects on glioblastoma multiforme (GBM) brain tumors and colon cancer. In vitro, GBM cells were treated with BP, and the effects on proliferation, cell cycle, and apoptosis were determined. In vivo, the human GBM tumor, DBTRG-05MG and RG2, the rat GBM tumor, were injected subcutaneously or intracerebrally with BP. BP increased the expression of cyclin kinase inhibitor, including p21 and p27, to decrease the phosphorylation of the Rb proteins, and down-regulated the cell cycle regulators, resulting in cell arrest and apoptosis at the G0/G1 phase. We also examined BP-induced changes in gene expression by microarray screening using human GBM brain tumor cells. Among these genes, Nur77 is particularly interesting because it plays an important role in the apoptotic processes in various tumor cell lines. BP was able to increase Nur77 mRNA and protein expression in a time-dependent manner. After the GBM 8401 cells were treated with BP, Nur77 translocated from the nucleus to the cytoplasm while the cytochrome c was released from the mitochondria, and caspase-3 became activated. Since BP has difficulty passing through the blood-brain barrier, we developed a local release system that incorporates BP into a biodegradable polyanhydride material, p(CPP-SA) (BP/Wafer), and investigated its antitumor effects. We used two xenograft animal models, F344 rats (for rat GBM) and nude mice (for human GBM), which were injected with RG2 and DBTRG-05MG cells, respectively, for tumor formation and subsequently treated subcutaneously with BP Wafers. In addition, to study the effect of the interstitial administration of BP against cranial brain tumors, BP/Wafers were surgically placed in FGF-SV40 transgenic mice. Our BP/Wafer significantly reduced tumor size and prolonged survival in a dose-dependent manner.

Non-Canonical Regulation of Type I Collagen through Promoter Binding of SOX2 and Its Contribution to Ameliorating Pulmonary Fibrosis by Butylidenephthalide

Pulmonary fibrosis is a fatal respiratory disease that gradually leads to dyspnea, mainly accompanied by excessive collagen production in the fibroblast and myofibroblast through mechanisms such as abnormal alveolar epithelial cells remodeling and stimulation of the extracellular matrix (ECM). Our results show that a small molecule, butylidenephthalide (BP), reduces type I collagen (COL1) expression in Transforming Growth Factor beta (TGF-β)-induced lung fibroblast without altering downstream pathways of TGF-β, such as Smad phosphorylation. Treatment of BP also reduces the expression of transcription factor Sex Determining Region Y-box 2 (SOX2), and the ectopic expression of SOX2 overcomes the inhibitory actions of BP on COL1 expression. We also found that serial deletion of the SOX2 binding site on 3′COL1 promoter results in a marked reduction in luciferase activity. Moreover, chromatin immunoprecipitation, which was found on the SOX2 binding site of the COL1 promoter, decreases in BP-treated cells. In an in vivo study using a bleomycin-induced pulmonary fibrosis C57BL/6 mice model, mice treated with BP displayed reduced lung fibrosis and collagen deposition, recovering in their pulmonary ventilation function. The reduction of SOX2 expression in BP-treated lung tissues is consistent with our findings in the fibroblast. This is the first report that reveals a non-canonical regulation of COL1 promoter via SOX2 binding, and contributes to the amelioration of pulmonary fibrosis by BP treatment.

The Role of Biomaterials in Implantation for Central Nervous System Injury

Permanent deficits that occur in memory, sensation, and cognition can result from central nervous system (CNS) trauma that causes dysfunction and/or unregulated CNS regeneration. Some therapeutic approaches are preferentially applied to the human body. Therefore, cell transplantation, one of the therapeutic strategies, may be used to benefit people. However, poor cell viability and low efficacy are the limitations to cell transplantation strategies. Biomaterials have been widely used in several fields (e.g., triggering cell differentiation, guiding cell migration, improving wound healing, and increasing tissue regeneration) by modulating their characteristics in chemistry, topography, and softness/stiffness for highly flexible application. We reviewed implanted biomaterials to investigate the roles and influences of physical/chemical properties on cell behaviors and applications. With their unique molecular features, biomaterials are delivered in several methods and mixed with transplanted cells, which assists in increasing postimplanted biological substance efficiency on cell survival, host responses, and functional recovery of animal models. Moreover, tracking the routes of these transplanted cells using biomaterials as labeling agents is crucial for addressing their location, distribution, activity, and viability. Here, we provide comprehensive comments and up-to-date research of the application of biomaterials.

Induced Pluripotent Stem Cells: A Powerful Neurodegenerative Disease Modeling Tool for Mechanism Study and Drug Discovery

Many neurodegenerative diseases are progressive, complex diseases without clear mechanisms or effective treatments. To study the mechanisms underlying these diseases and to develop treatment strategies, a reliable in vitro modeling system is critical. Induced pluripotent stem cells (iPSCs) have the ability to self-renew and possess the differentiation potential to become any kind of adult cell; thus, they may serve as a powerful material for disease modeling. Indeed, patient cell-derived iPSCs can differentiate into specific cell lineages that display the appropriate disease phenotypes and vulnerabilities. In this review, we highlight neuronal differentiation methods and the current development of iPSC-based neurodegenerative disease modeling tools for mechanism study and drug screening, with a discussion of the challenges and future inspiration for application.