In vitro and in vivo biocompatibility of decellularized cellulose scaffolds functionalized with chitosan and platelet rich plasma for tissue engineering applications.

This study describes the fabrication of cellulose scaffold (CS) and cellulose-chitosan (CS/CHI) scaffolds from the immature endosperm of Borassus flabellifer (Linn.) (BF) loaded with platelet rich plasma (PRP). Thus, developed scaffolds were evaluated for their physicochemical and mechanical behavior, growth factor release and biological performance. Additionally, in vivo response was assessed in a sub cutaneous rat model to study vascularization, host inflammatory response and macrophage polarization. The results of this study demonstrated that CS and CS/CHI scaffolds with PRP demonstrated favorable physiochemical and morphogical properties. The scaffold groups CS-PRP and CS/CHI-PRP were able to release growth factors in a well sustained manner under physiological conditions. The presence of PRP in cellulosic scaffolds did show significant differences in their behavior when investigated under in vitro studies, where the release of diverse cytokines improved the cellular proliferation and differentiation of osteoblasts. Finally, the PRP enriched scaffolds when studied under in vivo conditions showed increased angiogenesis and re-epithelialization with adequate collagen deposition and tissue remodeling. Our results suggest that besides the conventional carrier systems, this new-generation of plant-based cellulosic scaffolds with/without any modification can serve as a suitable carrier for PRP encapsulation and release, which can be used in numerous tissue regenerative therapies.

Decellularized natural 3D cellulose scaffold derived from Borassus flabellifer (Linn.) as extracellular matrix for tissue engineering applications.

In this study, Borassus flabellifer (Linn.) (BF) immature endosperm was decellularized to produce three dimensional (3D) cellulose scaffolds that can support mammalian 3D cell culture. To this regard, we first evaluated the chemical composition, nutritive profile and pharmacological activities of BF endosperm. The results demonstrated that the BF tissue represented a complex concoction of polysaccharides with intrinsic phyto-ingredients which provide excellent pharmacological properties. Furthermore cellulosic scaffolds (CS) obtained from BF was treated with chitosan to produce cellulose-chitosan (CS/CHI) hybrid scaffolds. The comparative investigation on both scaffolds exhibited adequate swelling with controlled porosity and pore-size distribution. The physiochemical characterization showed reduced biodegradation, improved thermal stability and enhanced compressive strength in CS/CHI group. Biological studies reported favorable adhesion and proliferation of fibroblasts with evident cellular penetration and colonization on the both scaffolds. Taken together, plant derived cellulosic scaffolds could be used as an alternative scaffolding material in regenerative medicine.

Hydroxyapatite nanophases augmented with selenium and manganese ions for bone regeneration: Physiochemical, microstructural and biological characterization.

Hydroxyapatite (HAP) nanopowders with different manganese (Mn) and selenium (Se) contents with Mn/Ca and Se/P molar ratio of 1 mol%, 2.5 mol% and 5 mol% were synthesized by wet-co-chemical precipitation method. The results revealed that with either Mn or Se doping, ion-substituted apatite phase was achieved with good crystallographic features. The combined evidence obtained from spectrometric techniques revealed that nanocrystalline HAP was effectively doped with Mn and Se ions, where Se in form of SeO32- replaced PO43- and Mn2+ replaced Ca2+. Mn and Se doped HAP samples exhibited rod-like and needle-like morphology with strong tendency to form agglomerates. HAP enriched with Mn and Se represented a strong antibacterial effect and also showed prominent blood compatibility. From the biocompatibility testing, it was evident that Mn and Se doped HAP augmented the osteoblasts adhesion, migration and proliferation in a dose-dependent manner. To conclude from this study, it is clearly evident that the doping amount of both Mn and Se ions can determine the size and morphology of the final HAP product. Therefore, Mn and Se HAP nanopowders with molar ratio less than 5 mol% without any heat treatment can provide good crystallographic features to HAP with satisfying micro-structural, thermal and biological properties.

Recent trends in natural polysaccharide based bioinks for multiscale 3D printing in tissue regeneration: A review.

Biofabrication by three-dimensional (3D) printing has been an attractive technology in harnessing the possibility to print anatomical shaped native tissues with controlled architecture and resolution. 3D printing offers the possibility to reproduce complex microarchitecture of native tissues by printing live cells in a layer by layer deposition to provide a biomimetic structural environment for tissue formation and host tissue integration. Plant based biomaterials derived from green and sustainable sources have represented to emulate native physicochemical and biological cues in order to direct specific cellular response and formation of new tissues through biomolecular recognition patterns. This comprehensive review aims to analyze and identify the most commonly used plant based bioinks for 3D printing applications. An overview on the role of different plant based biomaterial of terrestrial origin (Starch, Nanocellulose and Pectin) and marine origin (Ulvan, Alginate, Fucoidan, Agarose and Carrageenan) used for 3D printing applications are discussed elaborately. Furthermore, this review will also emphasis in the functional aspects of different 3D printers, appropriate printing material, merits and demerits of numerous plant based bioinks in developing 3D printed tissue-like constructs. Additionally, the underlying potential benefits, limitations and future perspectives of plant based bioinks for tissue engineering (TE) applications are also discussed.

Importance of crosslinking strategies in designing smart biomaterials for bone tissue engineering: A systematic review.

Biomaterials are of significant importance in biomedical applications as these biological macromolecules have moderately replaced classical tissue grafting techniques owing to its beneficial properties. Despite of its favourable advantages, poor mechanical and degradative properties of biomaterials are of great concern. To this regard, crosslinkers have emerged as a smart and promising tool to augment the biological functionality of biopolymers. Different crosslinkers have been extensively used in past decades to develop bone substitutes, but the implications of toxic response and adverse reactions are truly precarious after implantation. Traditional crosslinker like glutaraldehyde has been widely used in numerous bio-implants but the potential toxicity is largely being debated with many disproving views. As alternative, green chemicals, enzymatic and non-enzymatic chemicals, bi-functional epoxies, zero-length crosslinkers and physical crosslinkers have been introduced to achieve the desired properties of a bone substitute. In this review, systematic literature search was performed on PubMed database to identify the most commonly used crosslinkers for developing promising bone like materials. The relevant articles were identified, analysed and reviewed in this paper giving due importance to different crosslinking methodologies and comparing their effectiveness and efficacy in regard to material composition, scaffold production, crosslinker dosage, toxicity and immunogenicity. This review summarizes the recent developments in crosslinking mechanism with an emphasis placed on their ability to link proteins through bonding reactions. Finally, this study also covers the convergent and divergent methodologies of crosslinking strategies also giving special importance in retrieving the current limitations and future opportunities of crosslinking modalities in bone tissue engineering.

Bioactive compound 1,8-Cineole selectively induces G2/M arrest in A431 cells through the upregulation of the p53 signaling pathway and molecular docking studies.

BACKGROUND: Callistemon citrinus has been traditionally known for its medicinal property. Recently, our research group identified 1,8-Cineole, as one of the predominant compound present in the hexane extract (HE-C), whose leaves have potent anticancer activity. HYPOTHESIS/PURPOSE: The present study was designed to isolate 1,8-Cineole from Callistemon citrinus plant and to determine their role in anticancer effects in in vitro using skin carcinoma cells. Moreover, the molecular mechanism of apoptosis and molecular docking studies were also investigated. STUDY DESIGN/METHODS: In vitro cytotoxicity test was performed with HE-C fractionates 1F, 2F, and 3F against A431 and HaCaT cell lines. MTT and AB assay demonstrated that 1F was toxic to cancer cells with no adverse effect to non-malignant cells and it was subjected to 1H NMR, 13C NMR spectroscopy and further characterized by FTIR and GC-MS analysis. On the basis of spectroscopic data, the metabolite was confirmed as 1,8-Cineole. RESULTS: Based on the cytotoxicity results, the well-characterized metabolite 1,8-Cineole was investigated upon to understand the mechanism that caused cancer cell death. In this process, the changes in mitochondrial membrane potential (ΔΨm) were confirmed by Rh-123/DAPI staining; the ultra structure was observed by TEM and quantified by flow cytometric analysis. These results proved that the compound effectively induced the apoptosis and G2/M phase arrest in A431 cells by increasing the expression of p53 and that it was monitored by FACS. Further, the expression of apoptotic proteins, such as Bax/Bcl-2, Cyt-c, caspase-9, and caspase-3 was confirmed by western blot. The molecular docking simulations predicted the hydrophobic interaction between 1,8-cineole with Bcl-2 and PARP1 receptor. CONCLUSIONS: 1,8-Cineole is a potential candidate for skin carcinoma, which is possible by regulating the p53 apoptotic signaling pathway.

Evaluation of in vitro anticancer activity of 1,8-Cineole-containing n-hexane extract of Callistemon citrinus (Curtis) Skeels plant and its apoptotic potential.

Plants are the source of a variety of secondary metabolites, which are often used in the anticancer activity. Discovering new anticancer drug from herbal source is more important in both biological and pharmacological activities. Hence, the objective of this study is to identify the anticancer agent in Callistemon citrinus (Curtis) Skeels (CC) for the treatment of cancer. Very recently we have reported an increased antioxidant activity in the ethanolic and methanolic extracts (EE and ME) of CC but significantly reduced activity (rather increased cytotoxicity), in the n-hexane extract (HE). In this study, the cytotoxicity of all the three solvent extracts was tested against A431, MG-63 and HaCaT cell lines by MTT assay. Interestingly HE has showed increased anti-proliferative effect against the cancer cells but was resisted by non-malignant cells. HPLC and GC-MS analysis revealed the presence of 1,8-Cineole as a predominant compound in HE, the semi-purified bioactive extract. Henceforth, this would be called HE-C and be used for further analyses to understand its mode of action on induced apoptosis/necrosis. Alamar blue assay of HE-C showed cytotoxicity and change in morphological characteristics, which was confirmed by AO/EB staining using fluorescence microscopy, ultra-structural features of apoptosis using SEM and TEM. HE-C induced cell death was also detected by FACS using FITC-labelled Annexin-V and Propidium iodide. ROS generation was monitored using DCF-DA by flow cytometry. The overall results suggested that the selective extract (HE-C) containing 1,8-Cineole has shown potential anti-cancer activity in a dose-dependent manner, and cell death was induced through ROS-mediated apoptosis. Our findings provide an insight into the potential of 1,8-Cineole as a novel drug for killing cancer cells.