Chitosan-based hydrogels to induce neuronal differentiation of rat muscle-derived stem cells.

In this study, we used a chitosan hydrogel as a 3-dimensional substrate for the attachment, proliferation, and differentiation of rat muscle-derived stem cells (rMDSCs) in the presence of valproic acid (VA). Chitosan solutions containing glycerol phosphate disodium salt form a hydrogel at body temperature. The chitosan hydrogel exhibited a porous 3-dimensional network that allowed the culture medium to penetrate. The chitosan hydrogel acted as a suitable biocompatible substrate for the attachment and proliferation of rMDSCs. On chitosan hydrogel in the presence of VA, rMDSCs exhibited higher expression of the neural markers, neuron-specific enolase (NSE) and beta tubulin III (Tuj-1), the oligodendrocyte marker, oligodendrocyte transcription factor 2 (Olig-2), and the astrocyte marker, glial fibrillary acidic protein (GFAP) than those in the absence of VA. Our results suggest that rMDSCs on a chitosan hydrogel in the presence of VA can differentiate into cells with a neural-like phenotype.

In vivo efficacy of an intratumorally injected in situ-forming doxorubicin/poly(ethylene glycol)-b-polycaprolactone diblock copolymer.

The effectiveness of systemically administered anticancer treatments is limited by difficulties in achieving therapeutic doses within tumors, a problem that is complicated by dose-limiting side effects to normal tissue. This work examined injectable in situ-forming gels as a localized drug-delivery system. An MPEG-PCL (MP) solution containing doxorubicin (Dox) existed in an emulsion-sol state at room temperature and rapidly gelled in vitro and in vivo at body temperature. The release of Dox from Dox-loaded MP gels was sustained in vitro over 20 days after an initial burst, indicating that the MP gel acted as a drug depot. Dox-loaded MP gels exhibited remarkable in vitro anti-proliferative activities against B16F10 cancer cells. In vivo experiments employing B16F10 cancer cell xenograft-bearing mice showed that a single intratumoral injection of Dox-loaded MP gel inhibited the growth of tumors as effectively as repeated injections of free Dox, and more effectively than a single dose of free Dox, or saline or gel alone. Consistent with the observed suppression of tumor growth, intratumorally injected free Dox or Dox released from Dox-loaded MP gels caused apoptosis of tumor cells. The tumor biodistribution of free Dox after 1 day was ∼90%, which dropped to ∼15% after 4 days. The biodistribution of Dox following a single injection of Dox-loaded MP gel was also ∼90% on day 1, but remained at ∼13%, even after 15 days. Only a small amount of Dox was found in other organ tissues following intratumoral injection, implying fewer off-target side effects.

Potential induction of rat muscle-derived stem cells to neural-like cells by retinoic acid.

Several recent studies have demonstrated that stem cell differentiation can be generated by derivatives of retinoic acid. In this study we chose retinoic acid (RA) for inducing neural differentiation of rat muscle-derived stem cells (rMDSCs). rMDSCs were pre-induced with 10 ng/ml basic fibroblast growth factor (bFGF) and then treated with 2 µM RA. After stimulation, RA induced rMDSCs to have a neural-like morphology after 1-7 days of in vitro differentiation. In the results of immunocytochemistry, rMDSC treated with RA showed abundant positive cells against the neuronal markers neuronal-specific enolase (NSE) and tubulin-ßIII (Tuj1). Also, 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase)-positive cells were observed, indicating oligodendrocyte lineage cells. However, positive cells against glial fibrillary acidic protein (GFAP), marker of astrocytes, were not detected. The mRNA profile of these cells included higher expression of NSE compared with those of non-treated cells in real-time PCR. From the data in this work, we suggest that rMDSCs can trans-differentiate into a neural-like phenotype under the RA conditions.

In vivo biocompatibility study of electrospun chitosan microfiber for tissue engineering.

In this work, we examined the biocompatibility of electrospun chitosan microfibers as a scaffold. The chitosan microfibers showed a three-dimensional pore structure by SEM. The chitosan microfibers supported attachment and viability of rat muscle-derived stem cells (rMDSCs). Subcutaneous implantation of the chitosan microfibers demonstrated that implantation of rMDSCs containing chitosan microfibers induced lower host tissue responses with decreased macrophage accumulation than did the chitosan microfibers alone, probably due to the immunosuppression of the transplanted rMDSCs. Our results collectively show that chitosan microfibers could serve as a biocompatible in vivo scaffold for rMDSCs in rats.

Small intestine submucosa sponge for in vivo support of tissue-engineered bone formation in the presence of rat bone marrow stem cells.

The aim of the current study was to visualize new bone formed in vivo on a small intestine submucosa (SIS) sponge used as a tissue-engineered scaffold for the repair of damaged bone. The SIS sponge provided a three-dimensional pore structure, and supported good attachment and viability of rat bone marrow stem cells (rBMSCs). To examine bone regeneration, we prepared full-thickness bilateral bone defects in the rat crania, and then treated the defects with an implanted SIS sponge or PGA mesh without or with rBMSCs, or left the defects untreated. Bone defects were evaluated by micro-CT and histologically after 2 and 4 weeks. Micro-CT demonstrated a trend toward a decrease in bone void in both the SIS sponge and SIS sponge/rBMSCs groups compared to the control and PGA mesh groups. At 4 weeks, bone formation in defects containing SIS sponge/rBMSCs was significantly greater than in all other groups. A histological analysis after 2 and 4 weeks of implantation showed localized collagen and osteocalcin deposition on SIS sponges and SIS sponges with rBMSCs. These in vivo results indicate that the SIS sponge, implanted at bone-removal defects, facilitated bone regeneration.