Understanding the process of angiogenesis in regenerating earthworm.

Developing blood vessels from the existing vasculature is vital for the growth of the organism, as well as for systematic wound healing and the repair process. In this study, we investigated the role of angiogenesis during the regeneration process in the earthworm, Eudrilus eugeniae, animal model. Briefly, the morphological examination of blood vessels in juvenile and mature worms is documented, along with the development of new blood vessels in regenerating blastema. However, in vivo and in vitro experiments with juvenile worms revealed that geraniol retards blastemal regeneration growth with undeveloped blood vessels, as compared to the control. The results of qRT-PCR, western blotting, and immunohistochemistry confirmed a reduced expression of VEGFR2 and WNT5A in the day 3 regenerating blastema of geraniol-treated worms, as compared to the control. We conclude that geraniol acts as a potent natural inhibitor of angiogenesis, thereby retarding the regeneration process in earthworms. In addition, for studying angiogenesis and screening effective angiogenesis inhibitors as drug candidates, the earthworm is an ambient animal model system.

Foldscope microscope, an inexpensive alternative tool to conventional microscopy-Applications in research and education: A review.

Microscope is a device used for the visualization of tiny objects which are not visible to the naked eye. Traditional microscopes have been crucial for the advancement of contemporary science and medicine. Recent advancements in the field of microscopy have fueled its exponential growth rate. However, due to their expensive cost and complicated structure, modern microscopes remain inaccessible to the majority of the public. Nonetheless, the foldscope paper microscope has made it possible for anyone to explore and understand the world of microbes and organisms. In this review, we have listed foldscope-based research projects in various domains, as well as their key properties when compared to traditional research microscopes. In addition, we have briefly explored the impact of a foldscope microscope on public health, clinical diagnostics, forensic science, agriculture, basic science, developmental biology, and education. Moreover, the major drawbacks of paper microscopes and the current steps being taken to upgrade foldscope and its features are discussed in this review. Finally, we have concluded with our perspective that the microscope may be updated to imitate the advancement of a conventional microscope. RESEARCH HIGHLIGHTS: The foldscope, a low-cost instrument for studying the microscopic world. Foldscope applications were compared to conventional microscopes in many sectors. The foldscope microscope's existing limitations and potential prospects are highlighted.

Exploring the effect of UV-C radiation on earthworm and understanding its genomic integrity in the context of H2AX expression.

Maintaining genomic stability is inevitable for organism survival and it is challenged by mutagenic agents, which include ultraviolet (UV) radiation. Whenever DNA damage occurs, it is sensed by DNA-repairing proteins and thereby performing the DNA-repair mechanism. Specifically, in response to DNA damage, H2AX is a key protein involved in initiating the DNA-repair processes. In this present study, we investigate the effect of UV-C on earthworm, Perionyx excavatus and analyzed the DNA-damage response. Briefly, we expose the worms to different doses of UV-C and find that worms are highly sensitive to UV-C. As a primary response, earthworms produce coelomic fluid followed by autotomy. However, tissue inflammation followed by death is observed when we expose worm to increased doses of UV-C. In particular, UV-C promotes damages in skin layers and on the contrary, it mediates the chloragogen and epithelial outgrowth in intestinal tissues. Furthermore, UV-C promotes DNA damages followed by upregulation of H2AX on dose-dependent manner. Our finding confirms DNA damage caused by UV-C is directly proportional to the expression of H2AX. In short, we conclude that H2AX is present in the invertebrate earthworm, which plays an evolutionarily conserved role in DNA damage event as like that in higher animals.

Developmental stages of zebrafish (Danio rerio) embryos and toxicological studies using foldscope microscope.

Zebrafish (Danio rerio), is a well-established vertebrate animal model widely used in developmental biology and toxicological research. In the present study, foldscope is used as an innovative tool to study the developmental stages and toxicological analysis of the zebrafish embryos. Briefly, the developmental stages, such as zygote, cleavage, blastula, gastrula, segmentation, and pharyngula formation are observed and documented using simple foldscope. Toxicological parameters upon exposure to different concentration of ethanol extract of Curcuma longa and its lead compound, ar-turmerone along with rhodamine B (bio-coupler) on zebrafish embryos are analyzed upto 72 hr using foldscopes in live condition. The lethal endpoints, such as coagulation, lack of somite formation, non-detachment of tail, and lack of heartbeat are clearly monitored and documented using foldscope. Bio-evaluation of test compounds with the aid of foldscope confirms that the toxicity is directly proportional to the concentration. Our results conclude that, ethanol extract of C. longa, ar-turmerone and rhodamine B exposed embryos remains healthy up to 96, 48, and 24 µg concentrations, respectively. Embryos exposed to higher concentrations become coagulated, however normal physiological active movement of tail lashing and heartbeat are evident in lower concentration exposed embryos. Except coagulation, no other abnormalities are observed and interestingly, the hatching ability is not delayed, when compared with the control embryos. It is confirmed that the test compounds are not highly toxic to zebrafish embryos. Hence it can be used for further analysis, especially for studying the neural-regeneration and its neuronal development in zebrafish embryos.

Heat-inactivated coelomic fluid of the earthworm Perionyx excavatus is a possible alternative source for fetal bovine serum in animal cell culture.

Fetal Bovine Serum (FBS) is used as a major supplement in culturing animal cells under in vitro conditions. Due to ethical concern, high cost, biosafety, and geographical as well as batchwise result variations, it is important to reduce or replace the use of FBS in animal cell culture. The major objective of this work is to evaluate the feasibility of heat-inactivated coelomic fluid (HI-CF) of the earthworm, Perionyx excavatus as a possible alternative for FBS in animal cell culture experiments. The coelomic fluid (CF) was extruded from the earthworm using electric shock method and used for the experiments. Electric shock method is a simple non-invasive technique, which has no harmful effect on earthworms. Mouse primary fibroblast and HeLa cell lines were used in this study. Among HI-CF, autoclaved CF and crude CF, the supplement of medium with HI-CF shows positive results. The processed HI-CF (90°C for 5 min) at 10% supplement in cell culture medium promote maximum cell growth but cells need the initial support of FBS for the attachment to the culture flask. Microscopic observation and immunofluorescence assay with actin and lamin A confirm that the cellular and molecular morphology of the cells is maintained intact. The HI-CF of earthworm, P. excavatus has shown better cellular viability when compared with FBS and making it possible as an alternative supplement to minimize the use of FBS.

Understanding the Role of the Clitellum in the Regeneration Events of the Earthworm Eudrilus eugeniae.

Regeneration is a complex mechanism to restore lost or damaged body parts. In earthworms, regeneration capability varies among different species, and it is important to explore the mechanism behind the regeneration process. Interestingly, regeneration in earthworms is either dependent or independent of clitellum segments. In the present study, juvenile earthworms (Eudrilus eugeniae) were amputated at 3 different sites, namely the head, clitellum, and tail segments (at segments 10, 15, and 30, respectively), and their regeneration ability was documented using a foldscope. The amputated segments having the intact clitellum were able to heal the wounds and form the regenerative blastema. The smaller portions of the amputated segments (segments 1-10 and 1-15) without intact clitellum were unable to heal the wound, and death occurs within 12-24 h. The larger portions of the amputated segments (segments 15 and 30 to anus) without intact clitellum were able to heal the wound but lacked the regeneration capability. In control worms, alkaline phosphatase (ALP) signals were observed at the anterior tip, clitellum, and gut epithelium tissues, whereas, upon amputation, the enriched signals from the clitellum diminished, but profound signals were observed at the amputation site and regenerative blastema. Interestingly, on days 3 and 4, blastemal tips lacked ALP signals due to initiation of the differentiation process in the regeneration blastema. In summary, using a foldscope microscope, the role of the clitellum in the regeneration mechanism was indicated by ALP activity.