Yen-Lin Chen

Baculovirus transduction of rat articular chondrocytes: roles of cell cycle

We have previously demonstrated highly efficient baculovirus transduction of primary rat articular chondrocytes, thus implicating the possible applications of baculovirus in gene‐based cartilage tissue engineering. However, baculovirus‐mediated gene expression in the chondrocytes is transient.

Monascuspiloin Induces Apoptosis and Autophagic Cell Death in Human Prostate Cancer Cells via the Akt and AMPK Signaling Pathways

Monascus pigments have been reported to possess anticancer effects in various cancer cells; however, the molecular mechanisms of their anticancer properties remain largely unknown. Monascuspiloin is an analogue of the Monascus pigment monascin, and its anticancer growth activity against human prostate cancer cells was evaluated using in vitro and in vivo models. Monascuspiloin effectively inhibits the growth of both androgen-dependent LNCaP and androgen-independent PC-3 human prostate cancer cells. Monascuspiloin preferentially induces apoptosis in LNCaP cells by attenuating the PI3K/Akt/mTOR pathway. In androgen-independent PC-3 cells, monascuspiloin induces G2/M arrest and autophagic cell death by an AMPK-dependent pathway. Induction of autophagy in PC-3 cells further sensitizes cells to apoptosis induced by monascuspiloin. Monascuspiloin inhibits tumor growth in nude mice bearing PC-3 xenografts through induction of apoptosis and autophagy. This study is the first to demonstrate that monascuspiloin has therapeutic potential for the treatment of both androgen-dependent and -independent human prostate cancers.

Co-Conjugating Chondroitin-6-Sulfate/Dermatan Sulfate to Chitosan Scaffold Alters Chondrocyte Gene Expression and Signaling Profiles

Co‐conjugating chondroitin‐6‐sulfate (CSC) and dermatan sulfate (DS) to chitosan scaffolds improves chondrocyte differentiation and extracellular matrix (ECM) production. To further elucidate the cellular responses to CSC/DS conjugation, gene expression profiles for the rat chondrocytes cultured on the CSC/DS/chitosan and chitosan‐only scaffolds were compared by reverse‐transcription PCR (RT‐PCR) and quantitative real‐time RT‐PCR (qRT‐PCR). Our data unraveled that the CSC/DS/chitosan scaffold resulted in low‐level expression of collagen I, IIA and X and potentiated the aggrecan, collagen II (including collagen IIB) and TIMP3 expression, but downregulated the decorin expression. Therefore CSC/DS/chitosan scaffold maintained the chondrocyte differentiation while minimized de‐differentiation and hypertrophy. Furthermore, CSC/DS conjugation affected the expression of 11 genes implicated in 9 signaling pathways (as unveiled by cDNA microarray) and upregulated the expression of TGF‐β1, Sox9, BMP2, PTHrP and Ihh (as confirmed by qRT‐PCR). These data suggested that the CSC/DS/chitosan scaffold potentiated the TGF‐β and Hedgehog pathways, which activated the expression of PTHrP and its downstream Sox9. The signals were transduced to elevate the expression of aggrecan, collagen II and TIMP3, and contributed to the well‐differentiated chondrocyte phenotype. Altogether, this study for the first time elucidated the roles of GAGs‐conjugated biomaterials in matrix production and breakdown, cellular differentiation and signal transduction at the molecular levels. Biotechnol. Bioeng. 2008;101: 821–830.

Augmented biosynthesis of cadmium sulfide nanoparticles by genetically engineered Escherichia coli

Microorganisms can complex and sequester heavy metals, rendering them promising living factories for nanoparticle production. Glutathione (GSH) is pivotal in cadmium sulfide (CdS) nanoparticle formation in yeasts and its synthesis necessitates two enzymes: γ‐glutamylcysteine synthetase (γ‐GCS) and glutathione synthetase (GS). Hereby, we constructed two recombinant E. coli ABLE C strains to over‐express either γ‐GCS or GS and found that γ‐GCS over‐expression resulted in inclusion body formation and impaired cell physiology, whereas GS over‐expression yielded abundant soluble proteins and barely impeded cell growth. Upon exposure of the recombinant cells to cadmium chloride and sodium sulfide, GS over‐expression augmented GSH synthesis and ameliorated CdS nanoparticles formation. The resultant CdS nanoparticles resembled those from the wild‐type cells in size (2–5 nm) and wurtzite structures, yet differed in dispersibility and elemental composition. The maximum particle yield attained in the recombinant E. coli was ≈2.5 times that attained in the wild‐type cells and considerably exceeded that achieved in yeasts. These data implicated the potential of genetic engineering approach to enhancing CdS nanoparticle biosynthesis in bacteria. Additionally, E. coli‐based biosynthesis offers a more energy‐efficient and eco‐friendly method as opposed to chemical processes requiring high temperature and toxic solvents.

Baculovirus-mediated growth factor expression in dedifferentiated chondrocytes accelerates redifferentiation: effects of combinational transduction.

Transduction of partially dedifferentiated rabbit chondrocytes with a baculovirus (Bac-CB) expressing bone morphogenetic protein-2 (BMP-2) reverses dedifferentiation and enhances matrix production. Hereby we examined whether transduction with Bac-CB in combination with another baculovirus expressing transforming growth factor-beta1 (TGF-beta1) or insulin-like growth factor-1 (IGF-1) synergistically augmented chondrogenic differentiation. Passage 3 rabbit articular chondrocytes were transduced by different baculovirus combinations: single transduction with Bac-CB, cotransduction with Bac-CB and Bac-CT (expressing TGF-beta1), cotransduction with Bac-CB and Bac-CI (expressing IGF-1), and transduction with Bac-CB followed by repeated transduction with Bac-CT, Bac-CI, or Bac-CB 5 days later. Transduced cells were encapsulated into alginate beads for culture. Among these strategies, only cotransduction with Bac-CB and Bac-CT led to improved redifferentiation when compared with Bac-CB single transduction, as evidenced by the enhanced expression of aggrecan and collagen IIB (Col IIB), suppressed expression of Col I and Col X, emergence of chondrocyte-specific lacunae, and elevated deposition of matrix molecules. The cotransduction also accelerated the expression of Sox9, Col IIB, and aggrecan. In summary, baculovirus-mediated coexpression of TGF-beta1 and BMP-2 synergistically accelerates the chondrocyte redifferentiation process and improves the maintenance of chondrocyte phenotype and accumulation of cartilage-specific matrix molecules.

Monasnicotinates A-D, four new pyridine alkaloids from the fungal strain Monascus pilosus BCRC 38093.

Four new pyridine derivatives, monasnicotinates A–D (1–4) were isolated from the red yeast rice of Monascus pilosus BCRC 38093. Their structures were elucidated on the basis of physicochemical evidence, in-depth NMR spectroscopic analysis, and high-resolution mass spectrometry. Their inhibitory effects on NO production was also evaluated.

Baculovirus transduction of chondrocytes elicits interferon-α/β and suppresses transgene expression

Baculovirus is an effective vector for gene delivery into primary chondrocytes and repeated baculovirus transduction (i.e. supertransduction) appears to be promising for prolonging transgene expression, but how supertransduction may influence baculovirus‐mediated gene delivery is unknown.

Monascusazaphilones A–C, three new azaphilone analogues isolated from the fungus Monascus purpureus BCRC 38108

Investigation of the 95% EtOH extract of red yeast rice fermented with the pink mutant of the fungus Monascus purpureus BCRC 38108 led to the isolation of three new azaphilone derivatives, namely monascusazaphilones A–C (1–3), together with two known compounds. Compounds 1–3 were isolated from this species for the first time. Their structures were elucidated by 1-D and 2-D nuclear magnetic resonance spectroscopy together with HR–ESI–MS analysis and comparison of the spectroscopic data with those reported in the literatures. All isolates were evaluated for their inhibitory effects on nitric oxide (NO) production by macrophages. Among the isolates, compound 1 demonstrated stronger inhibition on NO production.

SECONDARY METABOLITES ISOLATED FROM THE FUNGUS MONASCUS KAOLIANG-FERMENTED RICE

A chemical study of the EtOAc-soluble ethanolic extract of Monascus kaoliang BCRC 31506 (Eurotiaceae) fermented rice led to the isolation of a new furan-3-one derivative along with 9 known compounds, monascin (2), ankaflavin (3), N-trans-feruloyltyramine (4), vanillic acid (5), 4-acetonyl-3,5-dimethoxy-p-quinol (6), (+)-s-eudesmol (7), trans-phytol (8), aesculetin dimethyl ether (9) and (-)-matairesinol (10). Structure 1 is proposed for monascuskaoliaone on the basis of spectroscopic evidence (NMR, UV, IR, and HRMS). None of the compounds exhibited significant scavenging activity toward DPPH radical in bioautography or spectroscopic assays.

Chemical constituents of red yeast rice fermented with the fungus Monascus pilosus

A chemical study on the n-BuOH-soluble fraction of the 95% EtOH extract of red yeast rice fermented with the fungus Monascus pilosus (Eurotiaceae) has resulted in the isolation of one new azaphilone derivative, designated as monapilosusazaphilone (1), together with nine known compounds, (+)-5,5′-dimethoxylariciresinol (2), (+)-lariciresinol (3), (+)-5-methoxylariciresinol (4), a mixture of 3β-hydroxylstigmast-5-en-7-one (5) and 3β-hydroxystigmasta-5,22-dien-7-one (6), 6β-hydroxystigmast-4-en-3-one (7), 6β-hydroxystigmasta-4,22-dien-3-one (8), N-cis-feruloylmethoxytyramine (9), and scopoletin (10). Their structures were elucidated by 1D and 2D-NMR spectroscopy together with HR-ESI-MS analysis, and comparison of the spectroscopic data with those reported for structurally related compounds.