Journal of Clinical and Diagnostic Research, ISSN - 0973 - 709X

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Dr Mohan Z Mani

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Believers Church Medical College,
Thiruvalla, Kerala
On Sep 2018




Prof. Somashekhar Nimbalkar

"Over the last few years, we have published our research regularly in Journal of Clinical and Diagnostic Research. Having published in more than 20 high impact journals over the last five years including several high impact ones and reviewing articles for even more journals across my fields of interest, we value our published work in JCDR for their high standards in publishing scientific articles. The ease of submission, the rapid reviews in under a month, the high quality of their reviewers and keen attention to the final process of proofs and publication, ensure that there are no mistakes in the final article. We have been asked clarifications on several occasions and have been happy to provide them and it exemplifies the commitment to quality of the team at JCDR."



Prof. Somashekhar Nimbalkar
Head, Department of Pediatrics, Pramukhswami Medical College, Karamsad
Chairman, Research Group, Charutar Arogya Mandal, Karamsad
National Joint Coordinator - Advanced IAP NNF NRP Program
Ex-Member, Governing Body, National Neonatology Forum, New Delhi
Ex-President - National Neonatology Forum Gujarat State Chapter
Department of Pediatrics, Pramukhswami Medical College, Karamsad, Anand, Gujarat.
On Sep 2018




Dr. Kalyani R

"Journal of Clinical and Diagnostic Research is at present a well-known Indian originated scientific journal which started with a humble beginning. I have been associated with this journal since many years. I appreciate the Editor, Dr. Hemant Jain, for his constant effort in bringing up this journal to the present status right from the scratch. The journal is multidisciplinary. It encourages in publishing the scientific articles from postgraduates and also the beginners who start their career. At the same time the journal also caters for the high quality articles from specialty and super-specialty researchers. Hence it provides a platform for the scientist and researchers to publish. The other aspect of it is, the readers get the information regarding the most recent developments in science which can be used for teaching, research, treating patients and to some extent take preventive measures against certain diseases. The journal is contributing immensely to the society at national and international level."



Dr Kalyani R
Professor and Head
Department of Pathology
Sri Devaraj Urs Medical College
Sri Devaraj Urs Academy of Higher Education and Research , Kolar, Karnataka
On Sep 2018




Dr. Saumya Navit

"As a peer-reviewed journal, the Journal of Clinical and Diagnostic Research provides an opportunity to researchers, scientists and budding professionals to explore the developments in the field of medicine and dentistry and their varied specialities, thus extending our view on biological diversities of living species in relation to medicine.
‘Knowledge is treasure of a wise man.’ The free access of this journal provides an immense scope of learning for the both the old and the young in field of medicine and dentistry as well. The multidisciplinary nature of the journal makes it a better platform to absorb all that is being researched and developed. The publication process is systematic and professional. Online submission, publication and peer reviewing makes it a user-friendly journal.
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I wish JCDR a great success and I hope that journal will soar higher with the passing time."



Dr Saumya Navit
Professor and Head
Department of Pediatric Dentistry
Saraswati Dental College
Lucknow
On Sep 2018




Dr. Arunava Biswas

"My sincere attachment with JCDR as an author as well as reviewer is a learning experience . Their systematic approach in publication of article in various categories is really praiseworthy.
Their prompt and timely response to review's query and the manner in which they have set the reviewing process helps in extracting the best possible scientific writings for publication.
It's a honour and pride to be a part of the JCDR team. My very best wishes to JCDR and hope it will sparkle up above the sky as a high indexed journal in near future."



Dr. Arunava Biswas
MD, DM (Clinical Pharmacology)
Assistant Professor
Department of Pharmacology
Calcutta National Medical College & Hospital , Kolkata




Dr. C.S. Ramesh Babu
" Journal of Clinical and Diagnostic Research (JCDR) is a multi-specialty medical and dental journal publishing high quality research articles in almost all branches of medicine. The quality of printing of figures and tables is excellent and comparable to any International journal. An added advantage is nominal publication charges and monthly issue of the journal and more chances of an article being accepted for publication. Moreover being a multi-specialty journal an article concerning a particular specialty has a wider reach of readers of other related specialties also. As an author and reviewer for several years I find this Journal most suitable and highly recommend this Journal."
Best regards,
C.S. Ramesh Babu,
Associate Professor of Anatomy,
Muzaffarnagar Medical College,
Muzaffarnagar.
On Aug 2018




Dr. Arundhathi. S
"Journal of Clinical and Diagnostic Research (JCDR) is a reputed peer reviewed journal and is constantly involved in publishing high quality research articles related to medicine. Its been a great pleasure to be associated with this esteemed journal as a reviewer and as an author for a couple of years. The editorial board consists of many dedicated and reputed experts as its members and they are doing an appreciable work in guiding budding researchers. JCDR is doing a commendable job in scientific research by promoting excellent quality research & review articles and case reports & series. The reviewers provide appropriate suggestions that improve the quality of articles. I strongly recommend my fraternity to encourage JCDR by contributing their valuable research work in this widely accepted, user friendly journal. I hope my collaboration with JCDR will continue for a long time".



Dr. Arundhathi. S
MBBS, MD (Pathology),
Sanjay Gandhi institute of trauma and orthopedics,
Bengaluru.
On Aug 2018




Dr. Mamta Gupta,
"It gives me great pleasure to be associated with JCDR, since last 2-3 years. Since then I have authored, co-authored and reviewed about 25 articles in JCDR. I thank JCDR for giving me an opportunity to improve my own skills as an author and a reviewer.
It 's a multispecialty journal, publishing high quality articles. It gives a platform to the authors to publish their research work which can be available for everyone across the globe to read. The best thing about JCDR is that the full articles of all medical specialties are available as pdf/html for reading free of cost or without institutional subscription, which is not there for other journals. For those who have problem in writing manuscript or do statistical work, JCDR comes for their rescue.
The journal has a monthly publication and the articles are published quite fast. In time compared to other journals. The on-line first publication is also a great advantage and facility to review one's own articles before going to print. The response to any query and permission if required, is quite fast; this is quite commendable. I have a very good experience about seeking quick permission for quoting a photograph (Fig.) from a JCDR article for my chapter authored in an E book. I never thought it would be so easy. No hassles.
Reviewing articles is no less a pain staking process and requires in depth perception, knowledge about the topic for review. It requires time and concentration, yet I enjoy doing it. The JCDR website especially for the reviewers is quite user friendly. My suggestions for improving the journal is, more strict review process, so that only high quality articles are published. I find a a good number of articles in Obst. Gynae, hence, a new journal for this specialty titled JCDR-OG can be started. May be a bimonthly or quarterly publication to begin with. Only selected articles should find a place in it.
An yearly reward for the best article authored can also incentivize the authors. Though the process of finding the best article will be not be very easy. I do not know how reviewing process can be improved. If an article is being reviewed by two reviewers, then opinion of one can be communicated to the other or the final opinion of the editor can be communicated to the reviewer if requested for. This will help one’s reviewing skills.
My best wishes to Dr. Hemant Jain and all the editorial staff of JCDR for their untiring efforts to bring out this journal. I strongly recommend medical fraternity to publish their valuable research work in this esteemed journal, JCDR".



Dr. Mamta Gupta
Consultant
(Ex HOD Obs &Gynae, Hindu Rao Hospital and associated NDMC Medical College, Delhi)
Aug 2018




Dr. Rajendra Kumar Ghritlaharey

"I wish to thank Dr. Hemant Jain, Editor-in-Chief Journal of Clinical and Diagnostic Research (JCDR), for asking me to write up few words.
Writing is the representation of language in a textual medium i e; into the words and sentences on paper. Quality medical manuscript writing in particular, demands not only a high-quality research, but also requires accurate and concise communication of findings and conclusions, with adherence to particular journal guidelines. In medical field whether working in teaching, private, or in corporate institution, everyone wants to excel in his / her own field and get recognised by making manuscripts publication.


Authors are the souls of any journal, and deserve much respect. To publish a journal manuscripts are needed from authors. Authors have a great responsibility for producing facts of their work in terms of number and results truthfully and an individual honesty is expected from authors in this regards. Both ways its true "No authors-No manuscripts-No journals" and "No journals–No manuscripts–No authors". Reviewing a manuscript is also a very responsible and important task of any peer-reviewed journal and to be taken seriously. It needs knowledge on the subject, sincerity, honesty and determination. Although the process of reviewing a manuscript is a time consuming task butit is expected to give one's best remarks within the time frame of the journal.
Salient features of the JCDR: It is a biomedical, multidisciplinary (including all medical and dental specialities), e-journal, with wide scope and extensive author support. At the same time, a free text of manuscript is available in HTML and PDF format. There is fast growing authorship and readership with JCDR as this can be judged by the number of articles published in it i e; in Feb 2007 of its first issue, it contained 5 articles only, and now in its recent volume published in April 2011, it contained 67 manuscripts. This e-journal is fulfilling the commitments and objectives sincerely, (as stated by Editor-in-chief in his preface to first edition) i e; to encourage physicians through the internet, especially from the developing countries who witness a spectrum of disease and acquire a wealth of knowledge to publish their experiences to benefit the medical community in patients care. I also feel that many of us have work of substance, newer ideas, adequate clinical materials but poor in medical writing and hesitation to submit the work and need help. JCDR provides authors help in this regards.
Timely publication of journal: Publication of manuscripts and bringing out the issue in time is one of the positive aspects of JCDR and is possible with strong support team in terms of peer reviewers, proof reading, language check, computer operators, etc. This is one of the great reasons for authors to submit their work with JCDR. Another best part of JCDR is "Online first Publications" facilities available for the authors. This facility not only provides the prompt publications of the manuscripts but at the same time also early availability of the manuscripts for the readers.
Indexation and online availability: Indexation transforms the journal in some sense from its local ownership to the worldwide professional community and to the public.JCDR is indexed with Embase & EMbiology, Google Scholar, Index Copernicus, Chemical Abstracts Service, Journal seek Database, Indian Science Abstracts, to name few of them. Manuscriptspublished in JCDR are available on major search engines ie; google, yahoo, msn.
In the era of fast growing newer technologies, and in computer and internet friendly environment the manuscripts preparation, submission, review, revision, etc and all can be done and checked with a click from all corer of the world, at any time. Of course there is always a scope for improvement in every field and none is perfect. To progress, one needs to identify the areas of one's weakness and to strengthen them.
It is well said that "happy beginning is half done" and it fits perfectly with JCDR. It has grown considerably and I feel it has already grown up from its infancy to adolescence, achieving the status of standard online e-journal form Indian continent since its inception in Feb 2007. This had been made possible due to the efforts and the hard work put in it. The way the JCDR is improving with every new volume, with good quality original manuscripts, makes it a quality journal for readers. I must thank and congratulate Dr Hemant Jain, Editor-in-Chief JCDR and his team for their sincere efforts, dedication, and determination for making JCDR a fast growing journal.
Every one of us: authors, reviewers, editors, and publisher are responsible for enhancing the stature of the journal. I wish for a great success for JCDR."



Thanking you
With sincere regards
Dr. Rajendra Kumar Ghritlaharey, M.S., M. Ch., FAIS
Associate Professor,
Department of Paediatric Surgery, Gandhi Medical College & Associated
Kamla Nehru & Hamidia Hospitals Bhopal, Madhya Pradesh 462 001 (India)
E-mail: drrajendrak1@rediffmail.com
On May 11,2011




Dr. Shankar P.R.

"On looking back through my Gmail archives after being requested by the journal to write a short editorial about my experiences of publishing with the Journal of Clinical and Diagnostic Research (JCDR), I came across an e-mail from Dr. Hemant Jain, Editor, in March 2007, which introduced the new electronic journal. The main features of the journal which were outlined in the e-mail were extensive author support, cash rewards, the peer review process, and other salient features of the journal.
Over a span of over four years, we (I and my colleagues) have published around 25 articles in the journal. In this editorial, I plan to briefly discuss my experiences of publishing with JCDR and the strengths of the journal and to finally address the areas for improvement.
My experiences of publishing with JCDR: Overall, my experiences of publishing withJCDR have been positive. The best point about the journal is that it responds to queries from the author. This may seem to be simple and not too much to ask for, but unfortunately, many journals in the subcontinent and from many developing countries do not respond or they respond with a long delay to the queries from the authors 1. The reasons could be many, including lack of optimal secretarial and other support. Another problem with many journals is the slowness of the review process. Editorial processing and peer review can take anywhere between a year to two years with some journals. Also, some journals do not keep the contributors informed about the progress of the review process. Due to the long review process, the articles can lose their relevance and topicality. A major benefit with JCDR is the timeliness and promptness of its response. In Dr Jain's e-mail which was sent to me in 2007, before the introduction of the Pre-publishing system, he had stated that he had received my submission and that he would get back to me within seven days and he did!
Most of the manuscripts are published within 3 to 4 months of their submission if they are found to be suitable after the review process. JCDR is published bimonthly and the accepted articles were usually published in the next issue. Recently, due to the increased volume of the submissions, the review process has become slower and it ?? Section can take from 4 to 6 months for the articles to be reviewed. The journal has an extensive author support system and it has recently introduced a paid expedited review process. The journal also mentions the average time for processing the manuscript under different submission systems - regular submission and expedited review.
Strengths of the journal: The journal has an online first facility in which the accepted manuscripts may be published on the website before being included in a regular issue of the journal. This cuts down the time between their acceptance and the publication. The journal is indexed in many databases, though not in PubMed. The editorial board should now take steps to index the journal in PubMed. The journal has a system of notifying readers through e-mail when a new issue is released. Also, the articles are available in both the HTML and the PDF formats. I especially like the new and colorful page format of the journal. Also, the access statistics of the articles are available. The prepublication and the manuscript tracking system are also helpful for the authors.
Areas for improvement: In certain cases, I felt that the peer review process of the manuscripts was not up to international standards and that it should be strengthened. Also, the number of manuscripts in an issue is high and it may be difficult for readers to go through all of them. The journal can consider tightening of the peer review process and increasing the quality standards for the acceptance of the manuscripts. I faced occasional problems with the online manuscript submission (Pre-publishing) system, which have to be addressed.
Overall, the publishing process with JCDR has been smooth, quick and relatively hassle free and I can recommend other authors to consider the journal as an outlet for their work."



Dr. P. Ravi Shankar
KIST Medical College, P.O. Box 14142, Kathmandu, Nepal.
E-mail: ravi.dr.shankar@gmail.com
On April 2011
Anuradha

Dear team JCDR, I would like to thank you for the very professional and polite service provided by everyone at JCDR. While i have been in the field of writing and editing for sometime, this has been my first attempt in publishing a scientific paper.Thank you for hand-holding me through the process.


Dr. Anuradha
E-mail: anuradha2nittur@gmail.com
On Jan 2020

Important Notice

Original article / research
Year : 2022 | Month : February | Volume : 16 | Issue : 2 | Page : XC05 - XC09 Full Version

Anticarcinogenic Efficacy of Fucoxanthin on HepG2 Cell Lines


Published: February 1, 2022 | DOI: https://doi.org/10.7860/JCDR/2022/49462.16007
P Sangavi, K Langeswaran, S Gowtham Kumar

1. Research Scholar, Department of Bioinformatics, Alagappa University, Karaikudi, Tamil Nadu, India. 2. Assistant Professor, Department of Bioinformatics, Alagappa University, Karaikudi, Tamil Nadu, India. 3. Assistant Professor, Department of Allied Health Science, Faculty of Allied Health Sciences, Chettinad Hospital and Research Institute, Chettiand Academy of Research, Chennai, Tamil Nadu, India.

Correspondence Address :
K Langeswaran,
Assistant Professor, Department of Bioinformatics, Alagappa University,
Karaikudi, Tamil Nadu, India.
E-mail: dr.langeswaran@gmail.com

Abstract

Introduction: Hepatocellular Carcinoma (HCC) is considered one of the deadliest cancers which grow rapidly worldwide. Fucoxanthin can be acquired from edible brown seaweeds, reported that fucoxanthin has numerous physiological functions and biological abilities, and various medicinal properties.

Aim: To evaluate the anticarcinogenic efficiency of fucoxanthin against the Human Hepatoma Cell Line (HepG2).

Materials and Methods: This in-vitro examination and research was carried from June 2018 to August 2018, at Alagappa University, Karaikudi, Tamil Nadu, India. Fucoxanthin at different concentrations was taken to demonstrate its antiproliferative potential and its cytotoxic effect. In this present examination, based on the outcome of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, 80 μM and 100 μM were chosen for further in-vitro studies. It was noted that remarkable leakage of Lactate Dehydrogenase (LDH) and a noteworthy decrease in the Glutathione (GSH) content of 80μ M and 100μM of fucoxanthin treated HepG2 cells. Under light and fluorescence microscopic examination, there was a significant reduction in cell growth and cell proliferation in the fucoxanthin-treated HepG2 cells. Data were presented as the mean±standard deviation. The one-way analysis of variance (ANOVA) and Tukey’s multiple comparison method were used to find the level of significance.

Results: The human HepG2 to be studied and analysed were divided into group I: control group; group II: comprising those cells treated with fucoxanthin in concentration of 80 μM; group III: comprising those cells treated with fucoxanthine in concentration of 100 μM. The results of the MTT assay put forward that fucoxanthin has the cytotoxic and antiproliferative potential against HepG2 further fucoxanthin significantly alter the marker enzymes level (p-value <0.05). The therapeutic efficiency of fucoxanthin might be due to the antioxidant effect of the bioactive compound. In this present investigation, the results of in-vitro studies, fucoxanthin strongly put forward that it has potential anticarcinogenic efficacy against the hepatic tumour (p-value <0.05).

Conclusion: The results of this analysis undoubtedly symbolise that fucoxanthin has antioxidant activity and anticarcinogenic efficacy against primary liver cancer HCC. Due to their therapeutic efficacy, may be considered as an excellent candidate against HCC.

Keywords

Antioxidant, Hepatic tumour, Hepatoma cell line, Microscopic examination, Primary liver cancer

HCC, most common primary liver malignancy and it is the most leading deadly tumour in the world. Studies reported that men are mostly affected by HCC when compared with women, especially people over the age of 50 years. Chronic hepatitis B and C virus are the leading causative factors for HCC (1). Primary HCC, called hepatoma and is one of the common liver malignancies. One of the most effective preventions of HCC is to prevent hepatic viral infection and cirrhosis because these are the important risk factors that are responsible for HCC. The pathogenesis of HCC is comprised of various hereditary/epigenetic variations and changes with many flagging pathways that lead to a known heterogeneity of the sickness biologic and clinical conduct (2).

Fucoxanthin, one of the natural medicines obtained from brown seaw-eed such as wakame (Undaria pinnatifida) and hijiki (Hijikia fusiformis) and it is also found in some dietary supplements. Some biological studies reported that fucoxanthin contains different biological benefits and some physiological functions (3). It has antitumour, antioxidant, anti-inflammatory, antiobesity, and antidiabetic properties. Fucoxanthin’s well known metabolites are fucoxanthinol and amarouciaxanthin. Fucoxanthinol was changed over into amaouciaxanthin A which was transcendently appeared in liver microsomes of mice and HepG2 cells (4). The anticancer impacts of fucoxanthin and fucoxanthinol are conjectured to be brought about by apoptosis enlistment and cell cycle arrest. The objectives of this investigation were to bring out the antioxidant potential, cytotoxic and antioncogenic effect of fucoxanthin. The target of this present study is to scrutiny the antitumourigenic potency of fucoxanthin against HepG2.

Material and Methods

The present study was an in-vitro investigation carried out at Alagappa University Karaikudi, Tamil Nadu, India, from June 2018 to August 2018.

Reagents: Chemical reagents were commercially acquired from Sigma Aldrich, St Louis, covering Dimethyl Sulfoxide (DMSO), Ethidium bromide (EtBr), and essential fucoxanthin in online mode. Culture medium such as RPMI (Roswell Park Memorial Institute)-1640, sodium pyruvate, penicillin, streptomycin, and bovine serum was procured from High Media Chemical Laboratory Pvt Ltd, Bangalore. Trypsin-Ethylene Diamine Tetra-acetic Acid (EDTA) was purchased from the same company at the door of the Chennai branch. In-vitro, culture plates and dishes were collected from Apex biotech ltd., Chennai, Tamil Nadu. Other necessary solvents immaculateness was acquired from Glaxo laboratory. Mumbai, India.

Preparation of drug: Fucoxanthin was diluted in Dimethylsulfoxide (DMSO) to the optimum concentration arranged in order with RPMI medium and placed at 4°C after purification used with syringe filter.

HepG2 cells maintenance and fucoxanthin treatment: Human HepG2 cells were used for this investigation. HepG2 cell lines were procured for this investigations from National Centre for Cell Sciences, Department of Biotechnology, Pune, India. Subsequently (5), deciphered those actions of antineoplastic were represented in monolayer cell culture. By achieved, the enormous quantity of replicate culture techniques has opened up the way and is evident for exploring several compounds (6). HepG2 is a cell line originating from the liver tissue of humans and grown using the monolayer culture method in the presence of RPMI-1640 medium. A monolayer culture was established under a controlled environment, humidified atmosphere, used with RPMI-1640 medium, and blended with heat-inactivated Bovine Serum along with few antibiotics such as penicillin 100 IU/mL, streptomycin 10 μg/mL, and 1 mM sodium pyruvate in the ratio of 5% CO2 at 37°C, Subsequently, after three days’ time duration, a reformed medium was observed and incubated, for another one day. The culture was trypsinised well used with 0.05% trypsin and EDTA (0.02%). Among them, 1×104 thickness cells were prioritised and chosen for the seeding process in each well plate for the MTT analysis. With a thickness of 0.5×106 cells were selected for LDH leakage assay. For subsequent investigations such as the GSH test and others, cells were incubated with or none fucoxanthin for a period of 24 hours.

MTT assay for cell viability: 3 (4, 5 dimethyl thiazol 2 yl) 2, 5 diphenyl tetrazolium bromide (MTT) procedure was executed as defined (7). The feasibility of the cells was measured by MTT assay, which is based on the lessening of MTT by the mitochondrial dehydrogenase of intact cells to a purple formazan product. Using an Enzyme Linked Immunosorbent Assay (ELISA) plate reader, the quantity of formazan was determined and the percentage of viable cells was calculated by measuring the absorbance at 540 nm using an ELISA plate reader.

Experimental setup: The HepG2 cell lines were divided into three groups. Group I was control, Group II & III cell lines were planned to be treated with fucoxanthin based on the results of MTT assay. Cells were placed in a standard humidity and atmosphere of 5% CO2 and 95% air for a period of 48 hours. Based on the cell viability and cytotoxic assay, 80 and 100 μM were chosen for further enzymatic analysis and microscopic observations. Group II were treated with fucoxanthine in concentration of 80 μM and Group III with concentration of 100 μM.

Assessment of enzyme marker: Estimation of LDH leakage was measured using Grivell and Berry method (7). The reduction rate of total Glutathione (GSH) was estimated by following the method outlined by Moron MS et al., (8).

Light microscopic study: Morphological alteration and vicissitudes that occurred due to the treatment of fucoxanthin for 24 hours were visualised by an inverted light microscope was attempted (9). Both control and fucoxanthin-treated HepG2 cells were examined to observe the abnormalities that occurred.

Fluorescent microscopic observation: Cells were stained with the fluorescent dye (Rhodamine Dye). After adequate treatment, cells were recovered carefully, washed well, and suspended in Phosphate buffered saline (PBS). Followed by the Whole Cell Stains (WCS) straining, a buffer solution, 100 μg/mL of phosphate buffer were added to dissolve and cells were tested (10). Variations in the ingestion of the dye indicated the survival of cells under fluorescent output. Results were analysed. The cytotoxic impacts of different synthetic concoctions and characteristic substances on threatening tumour cells in culture have been broadly considered as an essential screening for antitumour exercises (11).

Statistical Analysis

Data are presented as the mean±standard deviation (SD). The comparison of the means of different groups was achieved with the help of the Statistical Package for Social Sciences (SPSS) student’s version 12.5 and one-way analysis of variance (ANOVA) followed by Tukey’s multiple comparison method was used. Statistically significant values were compared between the control and drug-treated groups. The significance was considered at the level of p-value <0.05.

Results

In the present investigation, the chemotherapeutic adequacy of Fucoxanthin against human HepG2 was carried out. In this setting cytotoxicity, LDH enzyme activity, GSH estimation, and microscopic examinations were performed. The antiproliferative impact of fucoxanthin on HepG2 cells was tried by MTT strategy, which is dependable to identify the expansion of cells. (Table/Fig 1) display various concentrations of fucoxanthin (20, 40, 60, 80, and 100 μM) for 24 hours, which brought about diminished cell multiplication in a dose and time-dependent pattern when compared with control cells (p-value <0.05). Hence the responses were analysed by dividing human HepG2 cells into three groups. Group I as control while Group II of those cells treated with fucoxanthine of 80 μM and Group III of those cells treated with concentration of 100 μM.

The degrees of LDH discharged into the mechanism of control and fucoxanthin-treated HepG2 cells are exhibited in (Table/Fig 2) (p-value <0.05). From this figure, it was seen that marker enzyme LDH leakage was elevated after 24 hours introduction of 80 and 100 μM fucoxanthin. LDH elevation in drug-treated cells when contrast was done with the control confirms the cytotoxic nature of fucoxanthin (p-value <0.05). Each bar represents mean±SD of control and treated cells. It is outstanding that, GSH assumes a significant occupation in securing cells and cell segments against oxidative pressure and in detoxification. It is frequently discovered that GSH levels are expanded in the medication-safe disease cells when contrast was done with the medication induced cells. Hindrance of GSH amalgamation or balance of GSH stockpiles in tumours to lessen anticancer medications opposition may comprise a novel anticancer methodology (12).

The levels of GSH content in control and fucoxanthin treated HepG2 cells were exhibited in (Table/Fig 3). The critical depletion of GSH (p-value <0.05) was seen in fucoxanthin-treated HepG2 cells at 80 and 100 μM concentrations when contrast was done with the control cells.

Each bar represents the mean SD of control and treated cells:

a. Group I vs Group II and Group III
b. Group II vs Group III

Apoptosis is a hereditarily controlled reaction of a characteristic cell framework that is required for a basic harmony between cell expansion and cell demise in typical improvement and support of homeostasis of a living being. Numerous chemotherapeutic specialists smoother the development of changed or dangerous cells by actuating apoptosis (13). An enlistment of tumour cell apoptosis is one of the productive focuses for medicating advancement and has become a significant concentration in the investigation of disease treatment. Along these lines, the light and fluorescence minuscule and DNA fracture investigation was concentrated to decide the nearness of apoptosis in fucoxanthin-treated HepG2 cells. (Table/Fig 4) indicated the morphological changes of control and fucoxanthin-treated HepG2 cells at the concentration of 80 and 100 μM for 24 hours of treatment. In fucoxanthin treated HepG2 cells, the obliteration of monolayer was observed. It is not observed in HepG2 cells without fucoxanthin treatment. Control HepG2 demonstrated the growing and adjusted morphology of the cells with dense chromatin and their film. Fucoxanthin-treated cells display condensed chromatin and a shapeless nucleus. Dynamic auxiliary adjustments and a decrease of HepG2 cell populaces were seen in both concentrations of fucoxanthin. (Table/Fig 5) declared that the fluorescent microscopic image of control and 80 and 100 μM fucoxanthin treated HepG2 cells for 24 hours. Normal cores show chromatin with a composed structure, while apoptotic cores show exceptionally condensed chromatin in HepG2 cells. In the present current investigation, typical live cells were showed up in Rhodamine dye shading.

Discussion

Albeit customarily malignant growth has been battled with the typical armamentarium of chemotherapy and high portions of coordinated radiation, of late there has been more consideration committed to fighting disease through nutritive methods (14). Specifically, wide scopes of bioactive supplements have been discovered valuable in the physiological fight against malignant growth. HCC is the most widely recognised malignant growth around the world. It is related to poor endurance. HCC is a forceful tumour-related with dreary anticipation. Careful resection and liver transplantation are the two remedial medicines for HCC, yet are relevant to just a little extent of patients with the beginning phase of tumours (15). As of now, there is no successful fundamental chemotherapy for HCC, though elective treatment methodologies, for example, transcatheter blood vessel chemoembolisation, percutaneous intratumoural ethanol infusion, and radiofrequency removal are for the most part for vindication and are pertinent just to patients who have tumours confined in the liver. HCC is unmistakably a sickness for which elective remedial modalities are created. An intensive comprehension of the pathogenesis of HCC along these lines holds the guarantee of finding successful chemoprevention and curing malignant growth (16). Appraisal of the in-vitro cytotoxicity has as of late gotten well known as an essential screening strategy for assessing the antitumour exercises of different synthetics and common substances (17). To examine the anticancer action of fucoxanthin on cell feasibility, the LDH spillage test and estimation of GSH were acted in the HepG2 cell line. In the present examination, fucoxanthin fundamentally diminished cell suitability in a focus subordinate way. The concealment of cell expansion actuated by fucoxanthin might be because of the acceptance of cell demise. Along these lines, the inhibitory action of fucoxanthin is proof of in-vitro cytotoxicity. Late examinations recommend that the MTT measure is solid to evaluate cell reasonability.

MTT is a yellow water-soluble tetrazolium salt. Metabolically dynamic cells can change over the salt to water-insoluble dim blue formazan gems (18). Hence, the measure of formazan created is corresponding to the number of living cells. From the outcomes, it is gathered that the introduction of various convergence of fucoxanthin (20, 40, 60, 80, and 100 μM/ml) for 24 hours came about the decline of cell expansion in a portion subordinate way. Fucoxanthin about represses half-cell populace at the convergence of 80 and 100 μM for 24 hours when contrast was done with control. Along these lines, the inhibitory impact on HepG2 cells by fucoxanthin unequivocally demonstrates the counter expansion property of Fucoxanthin. Ongoing investigations recommend that LDH is an increasingly solid and progressively precise marker to ponder the cytotoxicity. Since harmed cells are divided totally throughout delayed hatching with poisonous substances (19).

In the present investigation, the LDH spillage expanded fundamentally in fucoxanthin treated HepG2 cells when contrast was done and the control cells. This outcome proposes that the hindrance might be because of the antitumour advertisers of the fucoxanthin. In this manner, it is proposed that the LDH spillage in HepG2 cells might be because of the cytotoxic nature of fucoxanthin and affirm its antitumour movement. Various laborers have called attention to those intracellular oxidative metabolites that assume a noteworthy job in the guideline of cell death. In this way, the estimation of GSH in medicating treated HepG2 cells establishes the auxiliary proof for programmed cell death.

Tripeptide GSH comprises a bizarre peptide connection between the amine gathering of cysteine and the carboxyl gathering of the glutamate side-chain. GSH, a cancer prevention agent that shields cells from responsive oxygen species, for example, free radicals and peroxides. GSH is nucleophilic at sulfur and assaults harmful electrophilic conjugate acceptors. GSH is originating only in its decreased structure, subsequently, the protein that returns it from its oxidised structure, GSH reductase, is constitutively dynamic and inducible upon oxidative pressure (20). The proportion of diminished GSH to oxidised GSH inside cells is utilised deductively as a proportion of cell lethality. Decreased GSH has been estimated to assume a job in the salvage of cells from apoptosis, by buffering an endogenously actuated oxidative pressure. The beginning of apoptosis was related to a fall of intracellular GSH in various cell frameworks. Loss of GSH was demonstrated to be firmly combined with various downstream occasions in apoptosis (21). It is realised that the lethality of antitumour medications may to a great extent rely upon the intracellular degree of decreased GSH.

GSH is known as a substrate in both conjugation responses and decreases responses, catalysed by GSH S-transferase compounds in the cytosol, microsomes, and mitochondria. Be that as it may, it is likewise fit for taking an interest in non enzymatic conjugation with certain synthetic concoctions. GSH exists in decreased (GSH) and oxidised (GSSG) states. In the decreased express, the thiol gathering of cysteine can give a lessening comparable (H++e-) to other temperamental particles, for example, receptive oxygen species. In giving an electron, GSH itself gets responsive, yet promptly responds with another receptive GSH to frame GSH disulfide (GSSG) and such a response is conceivable because of the generally high convergence of GSH in cells (up to 5 mM in the liver). GSH can be recovered from GSSG by the catalyst GSH reductase. This recommends Reactive Oxygen Species (ROS), particularly H2O2 may be associated with the procedure of apoptosis (22). Concentrates in an assortment of cell types recommend that disease chemotherapeutic medications actuate tumour cell apoptosis to a limited extent by expanding the development of ROS. Since, GSH is known to ensure cells against the danger of various operators.

In this current study, it was seen that the degrees of GSH were essentially diminished in fucoxanthin-treated HepG2 cells at the grouping of 80 and 100 μM. This demonstrates the lessening in GSH might be engaged with the restraint of HepG2 cell development and the basis of apoptosis. Consumption of GSH has been depicted for a few operators, for example, oxidative and alkylating specialists in different cell types. The present investigation demonstrates that the fucoxanthin may quickly initiate intracellular oxidation in HepG2 cells and leads to apoptosis which causes cell passing. It is theorised that fucoxanthin may incite ROS generation in HepG2 cells and in this way induce apoptosis. Not withstanding, ROS was not the immediate factor to cause apoptosis incited by the medication, however intracellular ROS may regulate the qualities associated with cell death. In the present examination, fucoxanthin prompted oxidative anxieties upstream of flagging occasions that may modify the professional and antiapoptotic balance in the HepG2 cells.

The phenolic mixes are having their cancer prevention agent impacts as well as expert oxidant activities under the in-vitro conditions (23). In this examination, it appeared to be conceivable that treatment with fucoxanthin in HepG2 cells exhaust the GSH levels and potentiates somewhat of oxidation acceptance, which exchanging the method of death employing apoptosis. Along these lines, the cytotoxic activity of this medication might be credited to its genius oxidant activity on the cells. This might have the option to represent the inconsistency in-vitro cytotoxicity and in-vivo antitumour exercises of fucoxanthin. Apoptosis is vitality subordinate, firmly managed and specific physiological procedure that oversees the expulsion of excessive or faulty cells. It happens in an ordinary biological manner. It can likewise be activated by various pathologies. In sound tissues, the fundamental job of cell death is to keep up the ideal quantity of cells in tissues and organs by expelling the excess, harmed, or practically anomalous cells (24).

The most transcendent morphologic highlights of apoptosis are film blebbing, cell shrinkage, and chromatin build-up. Apoptosis can be initiated by various boosts including some cell harming specialists and malignant growth treatment. In this association, light, fluorescence infinitesimal perception, DNA fracture and protein articulation in fucoxanthin treated HepG2 cells were contemplated. In the present examination, light tiny perception of fucoxanthin examined HepG2 cells at groupings of 80 and 100 μM after 24 hours of introduction demonstrated the run-of-the-mill morphological highlights of apoptosis in HepG2 cells. The structural variations watched were the decrease in cell volume, cell shrinkage, decrease in chromatin build-up, and development of cytoplasmic blebs. Be that as it may, the control HepG2 cells were observed with the higher intersection of the monolayer with no obliteration. It was seen that the fluorescence infinitesimal examination indicated apoptosis in fucoxanthin-treated HepG2 cells at convergences of 80 and 100 μM. In the present examination, control HepG2 cells and fucoxanthin-treated cells were recolored green in shading. When all is said in done, cytotoxic medications actuate an enormous breakage of DNA into oligonucleosomal parts is a late occasion of apoptosis. In this way, the fucoxanthin prompts DNA harm in HepG2 cells and along with these lines’ causes’ cell death. From this perception, it is deduced that fucoxanthin may apply an anticancer impact through DNA harm in HepG2 cells and advancing apoptosis. Hence, the likely utilisation of fucoxanthin and its derivatives, as co-adjuvant specialists in the therapy of disease should be added in-vivo and in-silico researched.

Limitation(s)

Combined therapies of fucoxanthin or fucoxanthinol with normal anticancer medicines can uphold regular restorative procedures by lessening drug opposition. Undoubtedly, as an anticancer atom, fucoxanthinol seems, by all accounts, to be a more viable bioactive compound than fucoxanthin. Hence, the likely utilisation of fucoxanthinol and other fucoxanthin subordinates, as co-adjuvant specialists in the therapy of disease ought to be additionally researched.

Conclusion

From the present analysis, it was concluded that fucoxanthin has notable cytotoxic and antiproliferative efficacy by damaging carcinogenic cells through realtering the cancer cell growth factors. It has apoptotic efficiency which was confirmed in microscopic investigated HepG2 cells. Previously, it has been reported that bioactive compounds flavonoids, alkaloids phenolics, and terpenoids exhibited antineoplastic activity in various in-vitro studies which supports these conclusions. Thus, it was powerfully recommended that fucoxanthin is a spectacular contender against human hepatoma cells HepG2.

Acknowledgement

The authors thankfully acknowledge the University Grants Commission (UGC), Government of India for their financial support to carry out the present research work. The authors also acknowledge the MHRD-RUSA 2.0 {F.24/51/2014-U, Policy (TNMulti-Gen), Department of Education, Government of India} for the laboratory facilities.

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DOI and Others

DOI: 10.7860/JCDR/2022/49462.16007

Date of Submission: Mar 16, 2021
Date of Peer Review: May 22, 2021
Date of Acceptance: Sep 30, 2021
Date of Publishing: Feb 01, 2022

AUTHOR DECLARATION:
• Financial or Other Competing Interests: Yes
• Was Ethics Committee Approval obtained for this study? NA
• Was informed consent obtained from the subjects involved in the study? NA
• For any images presented appropriate consent has been obtained from the subjects. NA

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