Development of gelatin/carboxymethyl chitosan/nano-hydroxyapatite composite 3D macroporous scaffold for bone tissue engineering applications

The present study delineates a relatively simpler approach for fabrication of a macroporous three-dimensional scaffold for bone tissue engineering. The novelty of the work is to obtain a scaffold with macroporosity (interconnected networks) through a combined approach of high stirring induced foaming of the gelatin/carboxymethyl chitosan (CMC)/nano-hydroxyapatite (nHAp) matrix followed by freeze drying. The fabricated macroporous (SGC) scaffold had a greater pore size, higher porosity, higher water retention capacity, slow and sustained enzymatic degradation rate along with higher compressive strength compared to that of non-macroporous (NGC, prepared by conventional freeze drying methodology) scaffold. The biological studies revealed the increased percentage of viability, proliferation, and differentiation as well as higher mineralization of differentiated human Wharton’s jelly MSC microtissue (wjhMSC-MT) on SGC as compared to NGC scaffold. RT-PCR also showed enhanced expression level of collagen type I, osteocalcin and Runx2 when seeded on SGC. μCT and histological analysis further revealed a penetration of cellular spheroid to a greater depth in SGC scaffold than NGC scaffold. Furthermore, the effect of cryopreservation on microtissue survival on the three-dimensional construct revealed significant higher viability upon revival in macroporous SGC scaffolds. These results together suggest that high stirring based macroporous scaffolds could have a potential application in bone tissue engineering

Environment Activatable Nanoprodrug: Two-Step Surveillance in the Anticancer Drug Release

Remedial cancer therapy deals with a large number of theranostic applications. However, systems, so far known, are only capable of single surveillance for both diagnostic and therapeutic modes of action. A nanosystem, which can be localized to the cancer and deliver the chemotherapeutic agent on demand, will provide effective therapeutic activity. Herein, we designed a single component nanoprodrug ANPD-X (Activatable Nano Pro-Drug-X) which indentified the tumor sites by fluorescent color change (signal 1, blue to green fluorescence) using H₂O₂-mediated oxidation of boronate fluorophore. In the next step, precise spatiotemporal irradiation of light only on identified tumor sites resulted in the release of anticancer drug chlorambucil. The real time information on drug release was achieved by a second fluorescence color change (signal 2, green to blue fluorescence). Thus, nanoprodrug ANPD-X provided overall two-step surveillance in the anticancer drug delivery. Activation of the ANPD-X after addition of H₂O₂ and drug release upon photoirradiation was investigated <i>in vitro</i> by monitoring its fluorescence in the HeLa cell line

Abrus Agglutinin, a type II ribosome inactivating protein inhibits Akt/PH domain to induce endoplasmic reticulum stress mediated autophagy-dependent cell death

Abrus agglutinin (AGG), a type II ribosome-inactivating protein has been found to induce mitochondrial apoptosis. In the present study, we documented that AGG-mediated Akt dephosphorylation led to ER stress resulting the induction of autophagy-dependent cell death through the canonical pathway in cervical cancer cells. Inhibition of autophagic death with 3-methyladenine (3-MA) and siRNA of Beclin-1 and ATG5 increased AGG-induced apoptosis. Further, inhibiting apoptosis by Z-DEVD-FMK and N-acetyl cysteine (NAC) increased autophagic cell death after AGG treatment, suggesting that AGG simultaneously induced autophagic and apoptotic death in HeLa cells. Additionally, it observed that AGG-induced autophagic cell death in Bax knock down (Bax-KD) and 5-FU resistant HeLa cells, confirming as an alternate cell killing pathway to apoptosis. At the molecular level, AGG-induced ER stress in PERK dependent pathway and inhibition of ER stress by salubrinal, eIF2 phosphatase inhibitor as well as siPERK reduced autophagic death in the presence of AGG. Further, our in silico and colocalization study showed that AGG interacted with pleckstrin homology (PH) domain of Akt to suppress its phosphorylation and consequent downstream mTOR dephosphorylation in HeLa cells. We showed that Akt overexpression could not augment GRP78 expression and reduced autophagic cell death by AGG as compared to pcDNA control, indicating Akt modulation was the upstream signal during AGG's ER stress mediated autophagic cell death. In conclusion, we established that AGG stimulated cell death by autophagy might be used as an alternative tumor suppressor mechanism in human cervical cancer. (c) 2016 Wiley Periodicals, Inc