Gelatin interpenetration in poly N-isopropylacrylamide network reduces the compressive modulus of the scaffold: A property employed to mimic hepatic matrix stiffness.

The progression of liver disease from normal to cirrhotic state is characterized by modulation of the stiffness of the extracellular matrix (ECM). Mimicking this modulation in vitro scaffold could provide a better insight into hepatic cell behavior. In this study, interpenetrating poly(N-isopropylacrylamide-co-gelatin) cryogels were synthesized in 48 different compositions to yield scaffolds of different properties. It was observed that a high concentration of N-isopropylacrylamide (NIPAAm) leads to the formation of small pores while gelatin interpenetration on poly-NIPAAm framework renders porous structure. Swelling properties and porosity of the gels decreased with an increase in NIPAAm concentration owing to the increased compactness of the gels. Gelatin interpenetration relaxed the gels and enhanced these properties. An increase in gelatin concentration led to a reduction in compressive moduli indicating that gelatin interpenetration in the poly-NIPAAm network softens the cryogel. With the increase in NIPAAm concentration, the effect of gelatin interpenetration in reducing the compressive moduli expanded. The cytocompatibility studies indicated that the gels are cell-adherent and compatible with HepG2. Furthermore, biochemical and real-time polymerase chain reaction studies revealed that HepG2 and Huh-7 cells cultured on scaffolds mimicking the ECM stiffness of normal liver (1.5-2.5 kPa) exhibited optimum liver-specific functionalities. Increasing the stiffness to fibrotic (4-9 kPa) and cirrhotic (10-20 kPa) ECM decreases the functionality.

Correction: Bioprospecting of fungal endophytes from Oroxylum indicum (L.) Kurz with antioxidant and cytotoxic activity.

[This corrects the article DOI: 10.1371/journal.pone.0264673.].

Unveiling the cytotoxic and anti-proliferative potential of green-synthesized silver nanoparticles mediated by Colletotrichum gloeosporioides.

Fungal endophytes are a putative source of bioactive metabolites that have found significant applications in nanomedicine due to their metabolic versatility. In the present study, an aqueous extract of the fungal endophyte, Colletotrichum gloeosporioides associated with a medicinal plant Oroxylum indicum, has been used for the fabrication of green silver nanoparticles (CgAgNPs) and further evaluated their cytotoxic and anti-proliferative activity. Bioanalytical techniques including UV-Vis spectral analysis revealed a sharp band at 435 nm and functional molecules from the aqueous extract involved in the synthesis of CgAgNPs were evidenced through FTIR. Further, the crystalline nature of CgAgNPs was determined through XRD analysis and microscopy techniques including AFM, TEM and FESEM demonstrated the spherical shape of CgAgNPs exhibiting a crystalline hexagonal lattice and the size was found to be in the range of 9-29 nm. The significant cytotoxic potential of CgAgNPs was observed against breast cancer cells, MDA-MB-231 and MCF-7 with IC50 values of 18.398 ± 0.376 and 38.587 ± 1.828 µg mL-1, respectively. The biochemical study revealed that the treatment of MDA-MB-231 and MCF-7 cells with CgAgNPs reduces glucose uptake, suppresses cell proliferation, and enhances LDH release, indicating reduced cell viability and progression. Moreover, our research revealed differential expression of genes associated with apoptosis, cell cycle inhibition and metastasis suppression, evidencing anti-proliferative activity of CgAgNPs. The main objective of the present study is to harness anti-breast cancer activity of novel biogenic nanoparticles synthesized using the aqueous extract of O. indicum associated C. gloeosporioides and study the underlying mechanistic pathway exerted by these mycogenic nanoparticles.

Ethyl Acetate Extract of Colletotrichum gloeosporioides Promotes Cytotoxicity and Apoptosis in Human Breast Cancer Cells.

Fungal endophytes are known to be a paragon for producing bioactive compounds with a variety of pharmacological importance. The current study aims to elucidate the molecular alterations induced by the bioactive compounds produced by the fungal endophyte Colletotrichum gloeosporioides in the tumor microenvironment of human breast cancer cells. GC/MS analysis of the ethyl acetate (EA) extract of C. gloeosporioides revealed the presence of bioactive compounds with anticancer activity. The EA extract of C. gloeosporioides exerted potential plasmid DNA protective activity against hydroxyl radicals of Fenton's reagent. The cytotoxic activity further revealed that MDA-MB-231 cells exhibit more sensitivity toward the EA extract of C. gloeosporioides as compared to MCF-7 cells, whereas non-toxic to non-cancerous HEK293T cells. Furthermore, the anticancer activity demonstrated by the EA extract of C. gloeosporioides was studied by assessing nuclear morphometric analysis and induction of apoptosis in MDA-MB-231 and MCF-7 cells. The EA extract of C. gloeosporioides causes the alteration in cellular and nuclear morphologies, chromatin condensation, long-term colony inhibition, and inhibition of cell migration and proliferation ability of MDA-MB-231 and MCF-7 cells. The study also revealed that the EA extract of C. gloeosporioides treated cells undergoes apoptosis by increased production of reactive oxygen species and significant deficit in mitochondrial membrane potential. Our study also showed that the EA extract of C. gloeosporioides causes upregulation of pro-apoptotic (BAX, PARP, CASPASE-8, and FADD), cell cycle arrest (P21), and tumor suppressor (P53) related genes. Additionally, the downregulation of antiapoptotic genes (BCL-2 and SURVIVIN) and increased Caspase-3 activity suggest the induction of apoptosis in the EA extract of C. gloeosporioides treated MDA-MB-231 and MCF-7 cells. Overall, our findings suggest that the bioactive compounds present in the EA extract of C. gloeosporioides promotes apoptosis by altering the genes related to the extrinsic as well as the intrinsic pathway. Further in vivo study in breast cancer models is required to validate the in vitro observations.

Assessment of Biological Activities of Fungal Endophytes Derived Bioactive Compounds Isolated from Amoora rohituka.

Fungal endophytes have remarkable potential to produce bioactive compounds with numerous pharmacological significance that are used in various disease management and human welfare. In the current study, a total of eight fungal endophytes were isolated from the leaf tissue of Amoora rohituka, and out of which ethyl acetate (EA) extract of Penicillium oxalicum was found to exhibit potential antioxidant activity against DPPH, nitric oxide, superoxide anion and hydroxyl free radicals with EC50 values of 178.30 ± 1.446, 75.79 ± 0.692, 169.28 ± 0.402 and 126.12 ± 0.636 µg/mL, respectively. The significant antioxidant activity of EA extract of P. oxalicum is validated through highest phenolic and flavonoid content, and the presence of unique bioactive components observed through high-performance thin layer chromatography (HPTLC) fingerprinting. Moreover, EA extract of P. oxalicum also displayed substantial anti-proliferative activity with IC50 values of 56.81 ± 0.617, 37.24 ± 1.26 and 260.627 ± 5.415 µg/mL against three cancer cells HuT-78, MDA-MB-231 and MCF-7, respectively. Furthermore, comparative HPTLC fingerprint analysis and antioxidant activity of P. oxalicum revealed that fungal endophyte P. oxalicum produces bioactive compounds in a host-dependent manner. Therefore, the present study signifies that fungal endophyte P. oxalicum associated with the leaf of A. rohituka could be a potential source of bioactive compounds with antioxidant and anticancer activity.

Bioprospecting of fungal endophytes from Oroxylum indicum (L.) Kurz with antioxidant and cytotoxic activity.

Oroxylum indicum (L.) Kurz, a medicinal plant, shows numerous pharmacological properties which may be attributed to the bioactive compounds produced by O. indicum or due to associated endophytes. In the present study, leaf of O. indicum was evaluated for the presence of associated fungal endophytes, and antioxidant and cytotoxic activities of bioactive compounds produced from them. Using culture-dependent approach, eight fungal endophytes belonging to five different genera were identified. Two endophytes Daldinia eschscholtzii and Ectophoma multirostrata have been reported for the first time from the leaf of O. indicum plant. High-performance thin-layer chromatography (HPTLC) of ethyl acetate (EA) extract of isolated fungal endophytes showed a distinct fingerprinting profile in EA extract of Colletotrichum gloeosporioides. Among identified endophytes, EA extract of C. gloeosporioides showed significant antioxidant activity against DPPH free radical, superoxide anion radical, nitric oxide radical and hydroxyl radical with EC50 values of 22.24±1.302 µg/mL, 67.46±0.576 µg/mL, 80.10±0.706 µg/mL and 61.55±1.360 µg/mL, respectively. EA extract of C. gloeosporioides exhibited potential cytotoxicity against HCT116, HeLa and HepG2 cancer cell lines with IC50 values of 76.59 µg/mL, 176.20 µg/mL and 1750.70 µg/mL, respectively. A comparative HPTLC fingerprinting and the antioxidant activity of C. gloeosporioides associated with two different hosts (leaf of O. indicum and dead twigs of other plant) showed that C. gloeosporioides produces bioactive compounds in a host-dependent manner.

Phytochemical Characterization, Antioxidant and Anti-Proliferative Properties of Rubia cordifolia L. Extracts Prepared with Improved Extraction Conditions.

Rubia cordifolia L. (Rubiaceae) is an important plant in Indian and Chinese medical systems. Extracts prepared from the root, stem and leaf have been used traditionally for the management of various diseases. Some of the known effects are anti-inflammation, neuroprotection, anti-proliferation, immunomodulation and anti-tumor. A comparative account of the extracts derived from different organs that lead to the identification of the most suitable solvent is lacking. We explored the presence of phytochemicals, antioxidant activity and anti-proliferative properties of a variety of solvent-based extracts of root, and methanol extracts of stem and leaf of R. cordifolia L. The antioxidant potential was determined by DPPH, hydrogen peroxide, nitric oxide and total antioxidant assays. The anti-proliferative nature was evaluated by MTT assay on HeLa, ME-180 and HepG2 cells. The composition of the extracts was determined by UPLC-UV-MS. We found that the root extracts had the presence of higher amounts of antioxidants over the stem and leaf extracts. The root extracts prepared in methanol exhibited the highest cytotoxicity in HepG2 cells. The main compounds identified through UPLC-UV-MS of the methanol extract give credibility to the previous results. Our comprehensive study corroborates the preference given to the root over the stem and leaf for extract preparation. In conclusion, we identified the methanol extract of the root to be the most suited to have bioactivity with anti-cancer potential.

A monoclonal antibody against annexin A2 targets stem and progenitor cell fractions in tumors.

The involvement of cancer stem cells (CSCs) in driving tumor dormancy and drug resistance is well established. Most therapeutic regimens however are ineffective in targeting these regenerative populations. We report the development and evaluation of a monoclonal antibody, mAb150, which targets the metastasis associated antigen, Annexin A2 (AnxA2) through recognition of a N-terminal epitope. Treatment with mAb150 potentiated re-entry of CSCs into the cell cycle that perturbed tumor dormancy and facilitated targeting of CSCs as was validated by in vitro and in vivo assays. Epigenetic potentiation further improved mAb150 efficacy in achieving total tumor regression by targeting regenerative populations to achieve tumor regression, specifically in high-grade serous ovarian adenocarcinoma.

Plant associated fungal endophytes as a source of natural bioactive compounds.

Endophytes are a potent source of bioactive compounds that mimic plant-based metabolites. The relationship of host plant and endophyte is significantly associated with alteration in fungal colonisation and the extraction of endophyte-derived bioactive compounds. Screening of fungal endophytes and their relationship with host plants is essential for the isolation of bioactive compounds. Numerous bioactive compounds with antioxidant, antimicrobial, anticancer, and immunomodulatory properties are known to be derived from fungal endophytes. Bioinformatics tools along with the latest techniques such as metabolomics, next-generation sequencing, and metagenomics multilocus sequence typing can potentially fill the gaps in fungal endophyte research. The current review article focuses on bioactive compounds derived from plant-associated fungal endophytes and their pharmacological importance. We conclude with the challenges and opportunities in the research area of fungal endophytes.

Recent technological advancements in stem cell research for targeted therapeutics.

Stem cells have characteristic features of self-renewal, pluripotency and differentiation, which are responsible for replenishment of tissue or organ. Stem cells are potentiated as therapeutic tool in drug targeting and regenerative medicine-from curing various neurological diseases and malignancies to congenital diseases. These technological advancements have established stem cells as future of medicine. However, due to ethico-social limitations, the use of embryonic stem cells (ESCs) has been avoided, while physiological availability of adult stem cells (ASCs) and induced pluripotent stem cells (iPSCs) has gained appropriate preference. These iPSCs are very much similar to ESCs in terms of their self-renewal and pluripotency. Here, we have summarized the technologies that have established stem cells isolation, their molecular marker and factors responsible for their maintenance. Different cellular (transcription factors, regulatory proteins, miRNA like miRNA-296, miRNA-145, etc.) and extracellular components transcend stem cell fate. Their identification and characterization involve development and efficient utilization of tools like magnetic activated cell sorting (MACS) and fluorescence activated cell sorting (FACS). Some of the technologies have been patented and spin-off's based on them have been commercialized. In conclusion, we present the future scope and possibilities that stem cell technologies behold for us. Graphical abstract Pictorial representation of therapeutic approaches for disease treatment using stem cell technology. Disease-specific adult stem cells are isolated along with niche cells by utilizing tools like FACS/MACS/LCM, etc. Thereafter, cells are reprogrammed through introduction of Yamanaka factors (Oct3/4, Sox2, c-myc, Klf4) to make induced pluripotent stem cell (iPSCs). The disease-specific iPSCs undergo genetic modification after delivery of therapeutic gene through retroviral vehicle. The genetically modified cells are introduced back in person with disease for therapeutic effects. FACS, fluorescence activated cell sorting; MACS, magnetic-activated cell sorting; LCM, laser capture microdissection; Oct3/4, octamer-binding transcription factor 3/4; Sox2, sex determining region Y-box 2; Klf4, Kruppel-like factor 4.

Clinical Stratification of High-Grade Ovarian Serous Carcinoma Using a Panel of Six Biomarkers.

Molecular stratification of high-grade serous ovarian carcinoma (HGSC) for targeted therapy is a pertinent approach in improving prognosis of this highly heterogeneous disease. Enabling the same necessitates identification of class-specific biomarkers and their robust detection in the clinic. We have earlier resolved three discrete molecular HGSC classes associated with distinct functional behavior based on their gene expression patterns, biological networks, and pathways. An important difference revealed was that Class 1 is likely to exhibit cooperative cell migration (CCM), Class 2 undergoes epithelial to mesenchymal transition (EMT), while Class 3 is possibly capable of both modes of migration. In the present study, we define clinical stratification of HGSC tumors through the establishment of standard operating procedures for immunohistochemistry and histochemistry based detection of a panel of biomarkers including TCF21, E-cadherin, PARP1, Slug, AnnexinA2, and hyaluronan. Further development and application of scoring guidelines based on expression of this panel in cell line-derived xenografts, commercial tissue microarrays, and patient tumors led to definitive stratification of samples. Biomarker expression was observed to vary significantly between primary and metastatic tumors suggesting class switching during disease progression. Another interesting feature in the study was of enhanced CCM-marker expression in tumors following disease progression and chemotherapy. These stratification principles and the new information thus generated is the first step towards class-specific personalized therapies in the disease.

Discrete molecular classes of ovarian cancer suggestive of unique mechanisms of transformation and metastases.

PURPOSE: Tumor heterogeneity and subsistence of high-grade serous ovarian adenocarcinoma (HGSC) classes can be speculated from clinical incidences suggesting passive tumor dissemination versus active invasion and metastases. EXPERIMENTAL DESIGN: We explored this theme toward tumor classification through two approaches of gene expression pattern clustering: (i) derivation of a core set of metastases-associated genes and (ii) resolution of independent weighted correlation networks. Further identification of appropriate cell and xenograft models was carried out for resolution of class-specific biologic functions. RESULTS: Both clustering approaches achieved resolution of three distinct tumor classes, two of which validated in other datasets. Networks of enriched gene modules defined biologic functions of quiescence, cell division-differentiation-lineage commitment, immune evasion, and cross-talk with niche factors. Although deviant from normal homeostatic mechanisms, these class-specific profiles are not totally random. Preliminary validation of these suggests that Class 1 tumors survive, metastasize in an epithelial-mesenchymal transition (EMT)-independent manner, and are associated with a p53 signature, aberrant differentiation, DNA damage, and genetic instability. These features supported by association of cell-specific markers, including PAX8, PEG3, and TCF21, led to the speculation of their origin being the fimbrial fallopian tube epithelium. On the other hand, Class 2 tumors activate extracellular matrix-EMT-driven invasion programs (Slug, SPARC, FN1, THBS2 expression), IFN signaling, and immune evasion, which are prospectively suggestive of ovarian surface epithelium associated wound healing mechanisms. Further validation of these etiologies could define a new therapeutic framework for disease management.