Search

The anterior cruciate ligament (ACL) is the most commonly injured intra-articular ligament of the knee, and limitations in existing reconstruction grafts have prompted an interest in tissue engineered solutions. Previously, we reported on a tissue-engineered ACL scaffold fabricated using a novel, three-dimensional braiding technology. A critical factor in determining cellular response to such a graft is material selection. The objective of this in vitro study was to optimize the braided scaffold, focusing on material composition and the identification of an appropriate polymer. The selection criteria are based on cellular response, construct degradation, and the associated mechanical properties. Three compositions of poly-alpha-hydroxyester fibers, namely polyglycolic acid (PGA), poly-L-lactic acid (PLLA), and polylactic-co-glycolic acid 82:18 (PLAGA) were examined. The effects of polymer composition on scaffold mechanical properties and degradation were evaluated in physiologically relevant solutions. Prior to culturing with primary rabbit ACL cells, scaffolds were pre-coated with fibronectin (Fn, PGA-Fn, PLAGA-Fn, PLLA-Fn), an important protein which is upregulated during ligament healing. Cell attachment and growth were examined as a function of time and polymer composition. While PGA scaffolds measured the highest tensile strength followed by PLLA and PLAGA, its rapid degradation in vitro resulted in matrix disruption and cell death over time. PLLA-based scaffolds maintained their structural integrity and exhibited superior mechanical properties over time. The response of ACL cells was found to be dependent on polymer composition, with the highest cell number measured on PLLA-Fn scaffolds. Surface modification of polymer scaffolds with Fn improved cell attachment efficiency and effected the long-term matrix production by ACL cells on PLLA and PLAGA scaffolds. Therefore based on the overall cellular response and its temporal mechanical and degradation properties in vitro, the PLLA braided scaffold pre-coated with Fn was found to be the most suitable substrate for ACL tissue engineering.

HTLV-1 oncoprotein Tax protein is known to be released from HTLV-1-transformed cells by a mechanism other than cell death; however, the mechanism of Tax secretion remains to be established. This study elucidates domains within Tax that contribute to its subcellular localization and secretion. Analysis of the amino acid sequence of Tax has revealed the presence of four putative secretory signals within the carboxy-terminal domain namely YTNI, LL, DHE and terminal V. Mutation of two putative signals (YTNI and DHE) resulted in aberrant subcellular localization of Tax, with cytoplasmic Tax accumulating in structures corresponding to the ER and Golgi. We have also studied the effect of these mutations on the secretion of Tax in baby hamster kidney cell (BHK-21) line. The results have demonstrated that mutating YTNI to ATNI resulted in approximately a 2-fold reduction in secretion. Mutation of LL to AA abrogated the intake of Tax, therefore, no secretion was observed. Mutating DHE and terminal V did not show any effect on Tax secretion, however, a combination of DHE mutation with YTNI mutation or AA mutation resulted in an altered intake and secretion of Tax. These studies substantiate a link between cytoplasmic Tax and its subsequent secretion and also indicate potential amino acid signals that might direct secretion of Tax. from 2005 International Meeting of The Institute of Human Virology Baltimore, USA, 29 August – 2 September 2005