ANTIPROLIFERATIVE AND APOPTOTIC ACTIVITY OF SULFATED POLYSACCHARIDE ISOLATED FROM HYPNEA VALENTIAE RED SEAWEED IN HUMAN SKIN MALIGNANT MELANOMA CELLS

Objective: Malignant melanoma is a highly metastatic cutaneous cancer. Deregulated apoptosis has been identified as a major cause of cancer drug resistance. The objective of the study is to evaluate antiproliferative activity of Hypnea Valentiae extract in human skin malignant melanoma (SK-MEL) cells. Methods: In this study, sulfated polysaccharide fraction was precipitated from aqueous extract obtained from H. valentiae. MTT assay was used to determine the cell viability of the crude sulfated polysaccharide against SK-MEL cells and normal L6 cell line (Rat skeletal muscle). Acridine orange (AO) and Ethidium bromide (EB) staining method was applied to study induction of apoptosis in SK-MEL cells. Results: Dose-dependent reduction in cell viability was observed with an IC50 of 30 μg/ml in SK-MEL cancer cells. The sulfated polysaccharide treated SK-MEL cells followed by AO, EB staining, showed typical early apoptotic, and late apoptotic morphological changes. Conclusion: The isolated crude sulfated polysaccharide from H. valentiae produced potent growth inhibition and induction of apoptosis in SK-MEL cells but caused no cytotoxicity in normal L6 skeletal muscle cells

Ascorbic acid attenuates ethanol induced apoptotic and oxidative response by blocking the Bax, Bcl2 and Caspase signaling pathways

Background: Research evidence has demonstrated that oxidative stress plays important etiological role in pathogenesis of alcoholic liver disease. The agents having antioxidant property plays a promising therapeutic intervention in ALD. In our present study we investigate the effect of ascorbic acid on ethanol induced liver injury and molecular mechanism of ethanol induced apoptosis.Methods: Wistar albino rats were randomly divided into 4 groups with 6 animals in each group control, ethanol treatment 40% (2ml/100gm), ethanol+ascorbic acid 100mg/kg b.w. intra-gastric gavage, ethanol+silymarin 100mg/kg b.w. intra-gastric gavage for 21 days. Statistical analysis was carried out using one-way ANOVA followed by Tukey multiple comparision test.Results: Ethanol induced hepatotoxicity is evidenced by increased level of liver marker enzymes (AST, ALT, ALP and LDH) and lipid peroxidation whereas the level of antioxidants (SOD, CAT, GSH, VIT C and E) was significantly decreased. Our results are further supported by histopathological examination which shows drastic changes in liver architecture. Hepatic Bax, Bcl-2, Caspase 3 and Caspase 9 proteins expressions were altered. On contrary treatment with ascorbic acid ameliorated the changes induced by ethanol and improved liver architecture. Conclusions: Ascorbic acid as an antioxidant protect the liver from ethanol induced oxidative damage and apoptosis.

Synthesis of 4-(benzyloxy)biphenyl under a multi-site phase-transfer catalyst along with ultrasonication – A kinetic study

In this present study, the synthesis of 4-(benzyloxy)biphenyl has been successfully carried out using 4-phenylphenol and benzyl chloride in the presence of ultrasonic power (40 kHz, 300W) along with synthesized multi - site phase - transfer catalyst viz., N1,N2-dibenzyl-N1,N1,N2,N2-tetramethylethane-1,2-diaminium dibromide (MPTC). The pseudo first-order kinetic equation has been applied to describe the organic reaction. The various kinetic parameters on the preparation of 4-(benzyloxy)biphenyl has been carried out under ultrasonic frequency condition and synthesized multi - site phase transfer catalyst. Sonication has resulted in better selectivity, high yield, avoids the use of the high temperature, side products and reduces the time consumption. The activation energy (Ea) has been calculated from the experimental results

Immuno-affinity Purification of Insect Cell Expressed Rabies Virus Glycoprotein using a Conformational Specific Monoclonal Antibody

.Rabies is a disease of nervous system and causes progressive encephalitis with fatal outcome. The conformation-dependent epitopes on the glycoprotein (G) of rabies virus (RV) is responsible for the induction of virus neutralizing antibodies which is ultimately required to get complete protection from viral challenge. Therefore, a suitable chromatography technique is necessary to purify the tag free recombinant rabies virus glycoprotein (rRVG) without altering its immunogenic epitopes. The present study was undertaken to purify the rRVG using a conformational specific anti-rabies virus glycoprotein (RVG) mAb, M5B4, which binds to the natively folded G. The mAb had shown a significant kinetic interaction with RVG. The mAb immobilized onto the NHS-activated Sepharose 4 fast flow™ was used for the purification of rRVG by immuno-affinity chromatography (IAC). The bound rRVG was eluted in IAC using 0.1M glycine with pH 2.5 and the identity of the purified protein was confirmed by MALDI-TOF. The IAC purified rRVG induced neutralizing antibody response and 83% of the immunized mice were protected against intra-cerebral rabies virus challenge. The results indicate that the mAb based IAC method can be an effective purification technique for tag free rRVG with significant level of purity, without compromising the protein’s immunogenic potential

Towards Upcycling Biomass-Derived Crosslinked Polymers with Light

Photodegradable, recyclable, and renewable, crosslinked polymers from bioresources show promise towards developing a sustainable strategy to address the issue of plastics degradability and recyclability. Photo processes are not widely exploited for upcycling polymers in spite of the potential to have spatial and temporal control of the degradation in addition to being a green process. In this report we highlight a methodology in which biomass-derived crosslinked polymers can be programmed to degrade at ≈300 nm with ≈60 % recovery of the monomer. The recovered monomer was recycled back to the crosslinked polymer

CAR T-Cell-Based gene therapy for cancers: new perspectives, challenges, and clinical developments

Chimeric antigen receptor (CAR)-T cell therapy is a progressive new pillar in immune cell therapy for cancer. It has yielded remarkable clinical responses in patients with B-cell leukemia or lymphoma. Unfortunately, many challenges remain to be addressed to overcome its ineffectiveness in the treatment of other hematological and solidtumor malignancies. The major hurdles of CAR T-cell therapy are the associated severe life-threatening toxicities such as cytokine release syndrome and limited anti-tumor efficacy. In this review, we briefly discuss cancer immunotherapy and the genetic engineering of T cells and, In detail, the current innovations in CAR T-cell strategies to improve efficacy in treating solid tumors and hematologic malignancies. Furthermore, we also discuss the current challenges in CAR T-cell therapy and new CAR T-cell-derived nanovesicle therapy. Finally, strategies to overcome the current clinical challenges associated with CAR T-cell therapy are included as well

A New Approach for Loading Anticancer Drugs Into Mesenchymal Stem Cell-Derived Exosome Mimetics for Cancer Therapy

Exosomes derived from mesenchymal stem cells (MSCs) have been evaluated for their potential to be used as drug delivery vehicles. Synthetically personalized exosome mimetics (EMs) could be the alternative vesicles for drug delivery. In this study, we aimed to isolate EMs from human MSCs. Cells were mixed with paclitaxel (PTX) and PTX-loaded EMs (PTX-MSC-EMs) were isolated and evaluated for their anticancer effects against breast cancer. EMs were isolated from human bone marrow-derived MSCs. MSCs (4 × 106 cells/mL) were mixed with or without PTX at different concentrations in phosphate-buffered saline (PBS) and serially extruded through 10-, 5-, and 1-μm polycarbonate membrane filters using a mini-extruder. MSCs were centrifuged to remove debris and the supernatant was filtered through a 0.22-μm filter, followed by ultracentrifugation to isolate EMs and drug-loaded EMs. EMs without encapsulated drug (MSC-EMs) and those with encapsulated PTX (PTX-MSC-EMs) were characterized by western blotting, nanoparticle tracking analysis (NTA), and transmission electron microscopy (TEM). The anticancer effects of MSC-EMs and PTX-MSC-EMs were assessed with breast cancer (MDA-MB-231) cells both in vitro and in vivo using optical imaging. EMs were isolated by the extrusion method and ultracentrifugation. The isolated vesicles were positive for membrane markers (ALIX and CD63) and negative for golgi (GM130) and endoplasmic (calnexin) marker proteins. NTA revealed the size of MSC-EM to be around 149 nm, while TEM confirmed its morphology. PTX-MSC-EMs significantly (p < 0.05) decreased the viability of MDA-MB-231 cells in vitro at increasing concentrations of EM. The in vivo tumor growth was significantly inhibited by PTX-MSC-EMs as compared to control and/or MSC-EMs. Thus, MSC-EMs were successfully isolated using simple procedures and drug-loaded MSC-EMs were shown to be therapeutically efficient for the treatment of breast cancer both in vitro and in vivo. MSC-EMs may be used as drug delivery vehicles for breast cancers

In Vivo Tracking of Chemokine Receptor CXCR4-Engineered Mesenchymal Stem Cell Migration by Optical Molecular Imaging

CXCR4, the stromal cell-derived factor-1 receptor, plays an important role in the migration of hematopoietic progenitor/stem cells to injured and inflamed areas. Noninvasive cell tracking methods could be useful for monitoring cell fate. Therefore, in this study, we evaluated the efficacy of an intravenous infusion of genetically engineered mesenchymal stem cells (MSCs) overexpressing CXC chemokine receptor 4 (CXCR4) to home to the tumor, by optical imaging. We constructed a retroviral vector containing CXCR with dual reporter genes, eGFP and Fluc2, under the control of an EF1α promoter (pBABE-EF1α-CXCR4-eGFP-IRES-Fluc2). We also developed an eGFP-Fluc2 construct in the Retro-X retroviral vector (Retro-X-eGFP-Fluc2). MSCs were transduced with retroviruses to generate CXCR4-overexpressing MSCs (MSC-CXCR4/Fluc2) and MSCs (MSC/Fluc2). CXCR4 mRNA and protein expression was confirmed by RT-PCR and Western blotting, respectively, and it was higher in MSC-CXCR4/Fluc2 than in naive MSCs. eGFP expression was confirmed by confocal microscopy. The transfected MSC-CXCR4/Fluc2 cells showed higher migratory capacity than naive MSCs observed in Transwell migration assay. The in vivo migration of CXCR4-overexpressing MSCs to MDAMB231/Rluc tumor model by BLI imaging was also confirmed. Intravenous delivery of genetically modified MSCs overexpressing CXCR4 with a Fluc2 reporter gene may be a useful, noninvasive BLI imaging tool for tracking cell fate

In vivo Non-invasive Imaging of Radio-Labeled Exosome-Mimetics Derived From Red Blood Cells in Mice

Exosomes are natural nano-sized membrane vesicles that have garnered recent interest owing to their potential as drug delivery vehicles. Though exosomes are effective drug carriers, their production and in vivo biodistribution are still not completely elucidated. We analyzed the production of exosome mimetics (EMs) from red blood cells (RBCs) and the radio-labeling of the RBC-EMs for in vivo imaging. Engineered EMs from RBCs were produced in large-scale by a one-step extrusion method, and further purified by density-gradient centrifugation. RBC-EMs were labeled with technetium-99m (99mTc). For non-invasive imaging, 99mTc (free) or 99mTc-RBC-EMs were injected in mice, and their biodistribution was analyzed by gamma camera imaging. Animals were sacrificed, and organs were collected for further biodistribution analysis. RBC-EMs have similar characteristics as the RBC exosomes but have a 130-fold higher production yield in terms of particle numbers. Radiochemical purity of 99mTc-RBC-EMs was almost 100% till 2 h reduced to 97% at 3 h. Radio-labeling did not affect the size and morphology of RBC-EMs. In contrast to free 99mTc, in vivo imaging of 99mTc-RBC-EMs in mice showed higher uptake in the liver and spleen, and no uptake in the thyroid. Ex vivo imaging confirmed the in vivo findings. Furthermore, fluorescent imaging confirmed the nuclear imaging findings. Immunofluorescent imaging revealed that the hepatic uptake of RBC-EMs was significantly mediated by kupffer cells (resident hepatic macrophages). Our results demonstrate a simple yet large-scale production method for a novel type of RBC-EMs, which can be effectively labeled with 99mTc, and feasibly monitored in vivo by nuclear imaging. The RBC-EMs may be used as in vivo drug delivery vehicles