Cancer stem cells CSCs (tumour-initiating cells) are responsible for cancer metastasis and recurrence associated with resistance to conventional chemotherapy. This study generated MBA MD231 3D cancer stem cells enriched spheroids in serum-free conditions and evaluated the influence of combined doxorubicin/thymoquinone-loaded cockle-shell-derived aragonite calcium carbonate nanoparticles. Single loaded drugs and free drugs were also evaluated. WST assay, sphere forming assay, ALDH activity analysis, Surface marker of CD44 and CD24 expression, apoptosis with Annexin V-PI kit, cell cycle analysis, morphological changes using a phase contrast light microscope, scanning electron microscopy, invasion assay and migration assay were carried out; The combination therapy showed enhanced apoptosis, reduction in ALDH activity and expression of CD44 and CD24 surface maker, reduction in cellular migration and invasion, inhibition of 3D sphere formation when compared to the free drugs and the single drug-loaded nanoparticle. Scanning electron microscopy showed poor spheroid formation, cell membrane blebbing, presence of cell shrinkage, distortion in the spheroid architecture; and the results from this study showed that combined drug-loaded cockle-shell-derived aragonite calcium carbonate nanoparticles can efficiently destroy the breast CSCs compared to single drug-loaded nanoparticle and a simple mixture of doxorubicin and thymoquinone.
Background: Combination chemotherapy of anticancer drugs is extensively being researched since it could reduce multidrug resistance and side effects as a result of lower dosage of each drug. In this study, we evaluated the effects of doxorubicin-loaded (Dox-ACNP), thymoquinone-loaded (TQ-ACNP) and a combined doxorubicin/thymoquinone-loaded cockle shell-derived aragonite calcium carbonate nanoparticles (Dox/TQ-ACNP) on breast cancer cell line and compared with their free drugs counterpart. Methods: Cell viability using MTT assay, apoptosis with Annexin V-PI kit, morphological changes using contrast light microscope, scanning electron microscope and transmission electron microscope, cell cycle analysis, invasion assay, and scratch assay were carried out. The cell viability was evaluated in breast cancer cell line (MDA MB231), normal breast cells (MDF10A) and normal fibroblast (3T3). Results: MDA MB231 IC50 dosages of drug-loaded nanoparticle were not toxic to the normal cells. The combination therapy showed enhanced apoptosis, reduction in cellular migration and invasion when compared to the single drug-loaded nanoparticle and the free drugs. Scanning electron microscope showed presence of cell shrinkage, cell membrane blebbing, while transmission electron microscope showed nuclear fragmentation, disruption of cell membrane, apoptotic bodies, and disruption of mitochondrial cistern. Conclusion: The results from this study showed that the combined drug-loaded cockle shell-derived aragonite calcium carbonate nanoparticles (Dox/TQ-ACNP) showed higher efficacy in breast cancer cells at lower dose of doxorubicin and thymoquinone.
Majority of the studies on the effect of chitin and chitosan on growth and carcass characteristics of broiler chickens has concentrated more on shrimp chitin and shrimp chitosan, and often with contradictory results. Therefore, the objective of this present study is to evaluate and compare the effect of dietary chitin and chitosan from cricket and shrimp on growth performance, carcass, and organ characteristics of broiler chickens. One hundred fifty-day-old male Cobb500 broiler chicks of similar average weight were randomly allotted into one of the five dietary treatments with three replicates. Treatment 1 (T1) chicks were fed basal diet only (control), treatment 2 and 3 (T2 and T3) chicks were given basal diet with 0.5 g/kg diet of cricket chitin and cricket chitosan, respectively, while treatment 4 and 5 (T4 and T5) chicks were served basal diet with 0.5 g/kg diet of shrimp chitin and shrimp chitosan respectively. No significant variation occurred between cricket chitin and shrimp chitin, although data on growth performance were higher in cricket chitin, but growth performance varied significantly between cricket chitosan and shrimp chitosan. This study revealed that cricket chitin at 0.5 g/kg significantly improved growth performance, carcass quality, and organ characteristics of broilers more than chitosan. Birds fed basal diet alone, although gained more weight, also accumulated more fat having the poorest feed conversion ratio (FCR) and the highest mortality. However, carcass of birds fed cricket chitin was the leanest and thus economically beneficial as they consumed the least amount of feed with the best FCR.
Chitin ranks next to cellulose as the most important bio-polysaccharide which can primarily be extracted from crustacean shells. However, the emergence of new areas of the application of chitin and its derivatives are on the increase and there is growing demand for new chitin sources. In this study, therefore, an attempt was made to extract chitin from the house cricket (Brachytrupes portentosus) by a chemical method. The physicochemical properties of chitin and chitosan extracted from crickets were compared with commercial chitin and chitosan extracted from shrimps, in terms of proximate analysis in particular, of their ash and moisture content. Also, infrared spectroscopy, x-ray diffraction (XRD), scanning electron microscopy and elemental analysis were conducted. The chitin and chitosan yield of the house cricket ranges over 4.3%-7.1% and 2.4%-5.8% respectively. Chitin and chitosan from crickets compares favourably with those extracted from shrimps, and were found to exhibit some similarities. The result shows that cricket and shrimp chitin and chitosan have the same degree of acetylation and degree of deacetylation of 108.1% and 80.5% respectively, following Fourier transform infrared spectroscopy. The characteristic XRD strong/sharp peaks of 9.4 and 19.4° for α-chitin are common for both cricket and shrimp chitin. The percentage ash content of chitin and chitosan extracted from B. portentosus is 1%, which is lower than that obtained from shrimp products. Therefore, cricket chitin and chitosan can be said to be of better quality and of purer form than commercially produced chitin and chitosan from shrimp. Based on the quality of the product, chitin and chitosan isolated from B. portentosus can replace commercial chitin and chitosan in terms of utilization and applications. Therefore, B. portentosus is a promising alternative source of chitin and chitosan.
Growth hormones (GH) alone does not explain the growth rate in the chicken as growth in an animal is multi-factorial. Normal morphology of the intestinal villus and crypt, with adequate regulation of intestinal nutrient transporters, is essential to a healthy gut. Nutrition plays a significant role in gut health management, but information on the effect of dietary chitin and chitosan on gut morphology, gene expression of nutrient transporter, and serum levels of GH in broiler chickens is scanty. Thus, this study aimed at evaluating the comparative effect of dietary chitin and chitosan from cricket and shrimp on the small intestinal morphology, relative gene expression of intestinal nutrient transporters and serum level of GH in the broiler. A total of 150 day-old male Cobb500 broiler chicks were randomly allotted to one of the five treatment groups (n = 30). Treatment 1 was fed basal diet only, treatments 2 to 5 were fed a basal diet with 0.5 g cricket chitin, cricket chitosan, shrimp chitin, and shrimp chitosan, respectively, per kg diet. At days 21 and 42, duodenal and jejunal samples were assessed for structural morphology and jejunum for the relative gene expression of PepT1, EAAT3, SGLT1, and SGLT5 using quantitative real-time PCR. Results bared that dietary cricket chitosan and shrimp chitosan significantly (P < 0.05) improved jejunal villus height and reduced crypt depth without improving the body weight (BW). The gut morphology of birds under cricket chitin was poor and significantly (P < 0.05) different from other treated groups. Both the dietary chitin and chitosan at day 21 and only dietary chitosan at day 42 significantly (P < 0.05) down-regulated the relative mRNA expression of PepT1, EAAT3, SGLT1, and SGLT5 of broiler chickens. Treated groups differ non-significantly at both phases, while cricket chitin numerically increased the relative expression of PepT1, EAAT3, and SGLT1. Therefore, the potential of cricket chitin to improve BW and to up-regulate nutrient transporters is worthy of further exploration.