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Ultra structural Studies of Mouse Liver in Castrated Subjects Treated with Grape Juice

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Research Article | DOI: https://doi.org/10.31579/2690-4861/082

Ultra structural Studies of Mouse Liver in Castrated Subjects Treated with Grape Juice

  • Homady M. H 1*
  • ALquraishi, L 2
  • Ubeid, M. H 3
  • Juma, A. S. M 1

1* College of Science, Department of Biomedical Sciences, Cihan University-Erbil, Kurdistan Region, Iraq.

2 College of Dentistry, Babylon University, Iraq.

3 College of Science, Department of General Biology, Cihan University-Erbil, Kurdistan Region, Iraq.

*Corresponding Author: Homady, M. H, 1College of Science, Department of Biomedical Sciences, Cihan University-Erbil, Kurdistan Region, Iraq.

Citation: Homady M. H, ALquraishi, L, Ubeid, M. H, Juma, A. S. M. (2021) Ultra structural Studies of Mouse Liver in Castrated Subjects Treated with Grape Juice. International Journal of Clinical Case Reports and Reviews. 6(3); DOI: 10.31579/2690-4861/082

Copyright: © 2021 Homady M. H, This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Received: 07 December 2020 | Accepted: 25 January 2021 | Published: 27 January 2021

Keywords: liver; castration; grape juice; kupffer cells; scanning electron microscope

Abstract

The present study was designed to evaluate the alterations in the liver tissues especially ITO and Kupffer cells of both castrated and castrated mice treated with10 µl/g of grape juice. The present study was conducted on forty five healthy males of Swiss albino mice, which were divided into 3 groups (N= 15 mice per each group). The first group was intact (control); the second one was castrated and the third group was castrated treated with 10 µl/g of grape juice. The ultra-structure sections of liver tissue from intact (control) group showed normal structure of both hepatocytes and sinusoids, with smooth external surfaces. Whereas ultra-sections from castrated group resulted in: degeneration of hepatocytes, deterioration of sinusoids; rough external surfaces, with aggregations of lipid droplets and white blood cells as compared with the control group. However treatment of castrated subject's with10µl/g of grape juice resulted in activation the regeneration processes in both hepatocytes and sinusoids, with smooth appearance of the external surfaces as compared with control group.

The present study concluded that surgical castration aggravated increased hepatic steatosis and increased inflammatory response by increased activation of Kupffer and Ito cells, these effects could be reserved by aggravated increased grape juice administration.

Introduction

The mammalian liver is the largest internal digestive organ, which is indispensable in many essential physiologic processes and vulnerable to be impaired by a wide variety of factors, such as toxins, microorganisms, metabolic products, circulatory materials and metabolism of carbohydrates, lipids and proteins. [1, 2, 3].

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in the world. It is present in 30% of the general adult population. In reality, NAFLD comprise as a spectrum of hepatic abnormalities that are observable in liver histological slides, from a simple intrahepatic accumulation of fat (steatosis or nonalcoholic fatty liver, NAFL) to various degrees of necrotic inflammation (NASH nonalcoholic steatohepatitis, NASH) [ 4, 5, 6]. Kupffer cells, the hepatic resident macrophages, represent the largest group of macrophages in the body and account for about 20–25% of non-parenchymal cells in the liver (intrasinusoidal cells)  [7, 8].  As the critical component of the innate immune system, Kupffer cells can be activated by various endogenous and exogenous stimuli, and play a key role in regulating the phenotype and function of neighboring parenchymal and non-parenchymal cells [9]. Hepatic stellate cells (HSCs) also called Ito cells, fat-storing cells, lipocytes, perisinusoidal cells and vitamin A-storing cells located in the space of Disse between hepatocytes and sinusoidal endothelial cells. These cells constitute approximately 10%of the total number of liver cells [10, 11, 12 and 13], and their cytoplasm is especially rich in lipid droplets and show long branched cytoplasmic processes that may reduce the lumen of sinusoid capillaries, in such a way modulating the liver sinusoidal blood flow. When the liver is damaged, the hepatic stellate cells change their shape and transform via a process named “activation” into the myofibroblast [14, 15 and 16]. All stages of NAFLD including cirrhosis and hepatocellular carcinomaassociated with hepatocyte steatosis [17].  Castration is a way of studying the consequences of extreme testosterone deficiency in animal models [18]. Castration promotes progression to steatohepatitis through activation of the ER (endoplasmic reticulum) stress pathway and enhancement of macrovesicular droplet.

Some medicinal plants may exert promising pharmacological properties and improve the effectiveness of conventional medications as complementary agents [19]. Vitis vinifera (Grape) are one of the most consumed fruits globally. It possesses a wide range of pharmacological activities due to its rich polyphenol ingredients most of which have been demonstrated to have therapeutic or health promoting properties [20, 21], among them, flavonoids are the most abundant and widely studied, recent studies have shown that the beneficial health effects promoted by consumption of grape and grape products are attributed to the unique mix of polyphenolic compounds. As the largest group of grape polyphenols, flavonoids are the main candidates considered to have biological properties, including but not limited to antioxidant, anti-inflammatory, anti-cancer antimicrobial, antiviral, cardioprotective, neuroprotective and hepatoprotective [22, 23, and 24]. Therefore, the aim of the present study was to determinethe effects of castration, and treatment of castrated subjects by using grape juice on both Ito and Kupffer cells by using Scanning Electron Microscope (SEM).

Materials and Methods

Swiss albino male mice weighting between (14-17) g., and aged (3weeks) were used in the present study, the mice were obtained from the Animal House, Faculty of Science/ University of Kufa.  Animals were kept in ventilated cages, with a temperature of (25±2Cº) at 12:12 h light, dark cycle was used balanced,  rodent food pellet and water were provided ad libitum [25, 26 and 27].  All experimental protocols using live animals were first reviewed, approved and accepted according to guidelines for the care and use of laboratory animals in biomedical research [28, 29 and 30]. A total number of 45 Swiss albino mice were used in the present study. Animals were divided into 5 groups (N=15), and the treatment was started at the age of 21 days for 6 weeks as:

Group I: Intact male mice received tap-water as control.

Group II: Castrated male mice received tap-water as (positive group).

Group III: Castrated male mice treated daily with (10µl/g) of grape juice,

The surgical castration method was done according to [31, 32]. Black grape (Vitis vinifera) obtained from local market (Baghdad, Iraq) 100g of grape was blundered by using a commercial blender without separating the seeds, and then it was filtered to remove the residue. The resulting extract (10 mls was stored in the refrigerator at 4˚C, and used after one hour. The extract was prepared according to [33]. A previous study documented that 10µl/g/day of grape juice extract was effective dose [34, 35, 36]. For this protocol, we used in the present study 10µl /g/day and was given daily as orally administered for six weeks.

Animals were sacrificed at the end of the experiments, with using ketamine and xylazine as anesthetic drugs to anesthetize the mice. The preparation procedures for Scanning Electron Microscopy has been described by [33]. Number of both Ito and Kupffer cells in cytoplasmic color in form of brown scale intensity was done according to [37].

Results

The present findings of electron micrograph of liver tissue from intact male mice (control) showed a normal structure ofboth hepatocytes and sinusoids with normal smooth external surface(Figures, 1, 2).

Figure (1): Scanning electron micrographs of liver tissue from intact male mice (control) showing normal structure of both hepatocytes (H) and sinusoids (S).
Figure (2): Scanning electron micrographs of liver tissue from intact male mice showing normal smooth external surface.

However the ultrastructure of liver tissue from castrated male mice

Showed degeneration processes of hepatocytes with deterioration of sinusoid (Figure 3) as well as aggregation of lipid droplets with rough external surfaces (Figure 4). Moreover, castration was also resulted in aggregations of white blood cells (Figure 5).

Figure (3): Scanning electron micrograph of liver tissue from castrated group showing degeneration of hepatocytes (H), with distortion of sinusoids (S)
Figure (4): Scanning electron micrograph of liver tissue from castrated group showing aggregation of lipids droplet with rough external surface arrow indicated  Lipid droplets
Figure (5): Scanning electron micrograph of liver tissue from castrated group showing an aggregations of white blood cells (WBC) arrow indicated white blood cellsThe ultrastructure of liver subject from castrated group treated with 10µl/g of grape juice (Figure, 6) showed regeneration processes in both hepatocytes and sinusoids, with smooth appearance of external surfaces (Figure 7).
Figure (6): Scanning electron micrograph of liver tissue from castrated group treated with 10µl/g grape juice showing regeneration of structures in both hepatocytes (H) and sinusoids(S)
Figure (7): Scanning electron micrograph of liver tissue from castrated group treated with 10µl/g of grape juice showing normal structure with smooth external surfaces.

Discussion

Scanning Electron Microscopical Results: The present results of the electron micrograph of liver tissue from intact (control) group showed a normal structure of hepatic lobules with smooth external surface. Whereas the results of castratedsubjects revealed, degeneration processes of hepatocytes and deterioration of sinusoid, with rough external surfaces, as well as aggregations of lipid droplets with concomitant increase in activation of immunity through aggregation of white blood cells. The interpretation of these findings may be attributed to hepatic changes as a results of castration that caused damaged or/may reductions in sinusoidal perfusion arise initially from the effects of hypertrophied hepatic parenchymal cells, swollen with accumulation of lipids withinand on their external surfaces. The extension in parenchymal cell plates caused narrowing and distortion the lumen of sinusoids as well as the intrasinusoidal lumen. These results are in agreement with the findings of [38, 39, 40], who stated that, hepatic steatosis altering the architecture of the sinusoidal network. Studied by [41, 42], demonstrated that the simple steatosis fatty livers may progress to steatohepatitis, and then to fibrosing steatohepatitis with the initiation of capillarization of the sinusoids. The present results are in concomitant with the findings of [43, 44], who concluded that the adhesion of leukocytes to the sinusoidal endothelium is followed by leukocyte infiltration into the hepatic parenchyma to form inflammatory foci. Many studied demonstrated that the deposition of collagen in the space of Diss accentuates the narrowing and distortion of the sinusoidal lumen, further restricting microvascular blood flow [45, 46, and 47]. Electron micrograph of liver tissue from castrated group treated with grape juice showed regeneration processes in both hepatocytes and sinusoids with smooth appearance of external surfaces. Such effects in the present study may be attributed to the protective effects of polyphenolic compounds which have multiple biological effects, including antioxidant, inhibiting lipid peroxidation activity, reduce the lipid load in the liver and reduce the swelling of hepatocytes that resulted in the dilation of spaces between them.

Conclusions: The present study demonstrated that impaired metabolic process related with surgical castration enhanced immune and inflammatory response, represented by activation of Kupffer and Ito cells which lead to hepatic apoptosis and contribute to increase nonalcoholic fatty liver disease. On the other hand the intra gastric application of Grape juice has dramatic effects to restore liver structure in castrated animals.

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Thank you very much for publishing my Research Article titled “Comparing Treatment Outcome Of Allergic Rhinitis Patients After Using Fluticasone Nasal Spray And Nasal Douching" in the Journal of Clinical Otorhinolaryngology. As Medical Professionals we are immensely benefited from study of various informative Articles and Papers published in this high quality Journal. I look forward to enriching my knowledge by regular study of the Journal and contribute my future work in the field of ENT through the Journal for use by the medical fraternity. The support from the Editorial office was excellent and very prompt. I also welcome the comments received from the readers of my Research Article.

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Dr Suramya Dhamija

Dear Erica Kelsey, Editorial Coordinator of Cancer Research and Cellular Therapeutics Our team is very satisfied with the processing of our paper by your journal. That was fast, efficient, rigorous, but without unnecessary complications. We appreciated the very short time between the submission of the paper and its publication on line on your site.

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Bruno Chauffert

I am very glad to say that the peer review process is very successful and fast and support from the Editorial Office. Therefore, I would like to continue our scientific relationship for a long time. And I especially thank you for your kindly attention towards my article. Have a good day!

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Baheci Selen

"We recently published an article entitled “Influence of beta-Cyclodextrins upon the Degradation of Carbofuran Derivatives under Alkaline Conditions" in the Journal of “Pesticides and Biofertilizers” to show that the cyclodextrins protect the carbamates increasing their half-life time in the presence of basic conditions This will be very helpful to understand carbofuran behaviour in the analytical, agro-environmental and food areas. We greatly appreciated the interaction with the editor and the editorial team; we were particularly well accompanied during the course of the revision process, since all various steps towards publication were short and without delay".

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Jesus Simal-Gandara

I would like to express my gratitude towards you process of article review and submission. I found this to be very fair and expedient. Your follow up has been excellent. I have many publications in national and international journal and your process has been one of the best so far. Keep up the great work.

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Douglas Miyazaki

We are grateful for this opportunity to provide a glowing recommendation to the Journal of Psychiatry and Psychotherapy. We found that the editorial team were very supportive, helpful, kept us abreast of timelines and over all very professional in nature. The peer review process was rigorous, efficient and constructive that really enhanced our article submission. The experience with this journal remains one of our best ever and we look forward to providing future submissions in the near future.

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Dr Griffith

I am very pleased to serve as EBM of the journal, I hope many years of my experience in stem cells can help the journal from one way or another. As we know, stem cells hold great potential for regenerative medicine, which are mostly used to promote the repair response of diseased, dysfunctional or injured tissue using stem cells or their derivatives. I think Stem Cell Research and Therapeutics International is a great platform to publish and share the understanding towards the biology and translational or clinical application of stem cells.

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Dr Tong Ming Liu

I would like to give my testimony in the support I have got by the peer review process and to support the editorial office where they were of asset to support young author like me to be encouraged to publish their work in your respected journal and globalize and share knowledge across the globe. I really give my great gratitude to your journal and the peer review including the editorial office.

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Husain Taha Radhi

I am delighted to publish our manuscript entitled "A Perspective on Cocaine Induced Stroke - Its Mechanisms and Management" in the Journal of Neuroscience and Neurological Surgery. The peer review process, support from the editorial office, and quality of the journal are excellent. The manuscripts published are of high quality and of excellent scientific value. I recommend this journal very much to colleagues.

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S Munshi

Dr.Tania Muñoz, My experience as researcher and author of a review article in The Journal Clinical Cardiology and Interventions has been very enriching and stimulating. The editorial team is excellent, performs its work with absolute responsibility and delivery. They are proactive, dynamic and receptive to all proposals. Supporting at all times the vast universe of authors who choose them as an option for publication. The team of review specialists, members of the editorial board, are brilliant professionals, with remarkable performance in medical research and scientific methodology. Together they form a frontline team that consolidates the JCCI as a magnificent option for the publication and review of high-level medical articles and broad collective interest. I am honored to be able to share my review article and open to receive all your comments.

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Tania Munoz

“The peer review process of JPMHC is quick and effective. Authors are benefited by good and professional reviewers with huge experience in the field of psychology and mental health. The support from the editorial office is very professional. People to contact to are friendly and happy to help and assist any query authors might have. Quality of the Journal is scientific and publishes ground-breaking research on mental health that is useful for other professionals in the field”.

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George Varvatsoulias

Dear editorial department: On behalf of our team, I hereby certify the reliability and superiority of the International Journal of Clinical Case Reports and Reviews in the peer review process, editorial support, and journal quality. Firstly, the peer review process of the International Journal of Clinical Case Reports and Reviews is rigorous, fair, transparent, fast, and of high quality. The editorial department invites experts from relevant fields as anonymous reviewers to review all submitted manuscripts. These experts have rich academic backgrounds and experience, and can accurately evaluate the academic quality, originality, and suitability of manuscripts. The editorial department is committed to ensuring the rigor of the peer review process, while also making every effort to ensure a fast review cycle to meet the needs of authors and the academic community. Secondly, the editorial team of the International Journal of Clinical Case Reports and Reviews is composed of a group of senior scholars and professionals with rich experience and professional knowledge in related fields. The editorial department is committed to assisting authors in improving their manuscripts, ensuring their academic accuracy, clarity, and completeness. Editors actively collaborate with authors, providing useful suggestions and feedback to promote the improvement and development of the manuscript. We believe that the support of the editorial department is one of the key factors in ensuring the quality of the journal. Finally, the International Journal of Clinical Case Reports and Reviews is renowned for its high- quality articles and strict academic standards. The editorial department is committed to publishing innovative and academically valuable research results to promote the development and progress of related fields. The International Journal of Clinical Case Reports and Reviews is reasonably priced and ensures excellent service and quality ratio, allowing authors to obtain high-level academic publishing opportunities in an affordable manner. I hereby solemnly declare that the International Journal of Clinical Case Reports and Reviews has a high level of credibility and superiority in terms of peer review process, editorial support, reasonable fees, and journal quality. Sincerely, Rui Tao.

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Rui Tao

Clinical Cardiology and Cardiovascular Interventions I testity the covering of the peer review process, support from the editorial office, and quality of the journal.

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Khurram Arshad

Clinical Cardiology and Cardiovascular Interventions, we deeply appreciate the interest shown in our work and its publication. It has been a true pleasure to collaborate with you. The peer review process, as well as the support provided by the editorial office, have been exceptional, and the quality of the journal is very high, which was a determining factor in our decision to publish with you.

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Gomez Barriga Maria Dolores

The peer reviewers process is quick and effective, the supports from editorial office is excellent, the quality of journal is high. I would like to collabroate with Internatioanl journal of Clinical Case Reports and Reviews journal clinically in the future time.

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Lin Shaw Chin

Clinical Cardiology and Cardiovascular Interventions, I would like to express my sincerest gratitude for the trust placed in our team for the publication in your journal. It has been a true pleasure to collaborate with you on this project. I am pleased to inform you that both the peer review process and the attention from the editorial coordination have been excellent. Your team has worked with dedication and professionalism to ensure that your publication meets the highest standards of quality. We are confident that this collaboration will result in mutual success, and we are eager to see the fruits of this shared effort.

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Maria Dolores Gomez Barriga