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

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Dr Mohan Z Mani

"Thank you very much for having published my article in record time.I would like to compliment you and your entire staff for your promptness, courtesy, and willingness to be customer friendly, which is quite unusual.I was given your reference by a colleague in pathology,and was able to directly phone your editorial office for clarifications.I would particularly like to thank the publication managers and the Assistant Editor who were following up my article. I would also like to thank you for adjusting the money I paid initially into payment for my modified article,and refunding the balance.
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Dr Mohan Z Mani,
Professor & Head,
Department of Dermatolgy,
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.
As an experienced dentist and an academician, I proudly recommend this journal to the dental fraternity as a good quality open access platform for rapid communication of their cutting-edge research progress and discovery.
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 : 2024 | Month : May | Volume : 18 | Issue : 5 | Page : EC06 - EC11 Full Version

Clinicopathological and Immunohistochemical Association of CD55 and CD59 in Colorectal Carcinoma: A Cross-sectional Study


Published: May 1, 2024 | DOI: https://doi.org/10.7860/JCDR/2024/69311.19379
Rathin Hazra, Ananya Khan, Rajib Kumar Mondal, Sarbari Kar Rakshit

1. Associate Professor, Department of Pathology, Diamond Harbour Government Medical College, Diamond Harbour, West Bengal, India. 2. Senior Resident, Department of Pathology, Nil Ratan Sircar Medical College, Kolkata, West Bengal, India. 3. Associate Professor, Department of Pathology, Nil Ratan Sircar Medical College, Kolkata, West Bengal, India. 4. Assistant Professor, Department of Pathology, Nil Ratan Sircar Medical College, Kolkata, West Bengal, India.

Correspondence Address :
Dr. Rathin Hazra,
Village Hasanpur, Town Joynagar, Joynagar Majilpur Municipality, Ward Number-13, Block-Jaynagar 1, PO-Ramakanta Nagar, PS-Jaynagar, Dist-South 24 Parganas, Jaynagar-74339, West Bengal, India.
E-mail: hazra_rathin@rediffmail.com

Abstract

Introduction: Colorectal Carcinoma (CRC) is one of the significant causes of fatality worldwide. High-grade and high-stage cancers contribute to this fatality. In most terminal cases, new immunomarkers (CD55 and CD59) are commonly detected as positive. The expression of these immunomarkers and their clinical relevance in CRC has not yet been firmly established. While their upregulation has been demonstrated in some CRC several groups have also reported that they are not highly expressed in CRC tissues.

Aim: To study the expression of CD55 and CD59 in CRC.

Materials and Methods: This was a cross-sectional study involving 90 patients with colorectal growth, conducted from February 2020 to July 2021 in the Department of Pathology in collaboration with the Department of General Surgery at Nil Ratan Sircar Medical College, Kolkata, West Bengal, India. Histopathological findings were evaluated, and associations of grading, staging, tumour type, tumour location, age, and sex were studied with immunomarkers (CD55 and CD59). Clinical parameters of the patients with CRC diagnosed by colonoscopic biopsy were also studied, including age, sex, clinical pictures, food habits, family history of CRC, and prior chemotherapy. The Chi-square test was used to determine the significance of the study. Data were analysed using the Statistical Package for the Social Sciences (SPSS) version 27.0 (IBM, Illinois, US). A p-value of ≤0.05 was considered statistically significant.

Results: A total of 90 CRC patients were included in this study, with the majority being male 56 (62.2%) aged 51-60 years. Most cases 59 (65.6%) were moderately differentiated adenocarcinoma, followed by poorly differentiated adenocarcinoma 18 (20%) and well-differentiated adenocarcinoma 13 (14.4%). About 62 (68.9%) and 28 (31.1%) patients showed strong and weak CD55 expression, respectively. In total, 65 (72.2%) and 25 (27.8%) patients exhibited strong and weak CD59 expression, respectively. The association of CRC grade and stage with CD55 and CD59 was significant (p-value=0.0001, p-value=0.0013, p-value=0.0001, and p-value=0.0001, respectively).

Conclusion: There are several variables to consider during the histopathological reporting of CRC, with tumour differentiation, grade, and stage being among the most important. Particularly, high-grade (poorly differentiated) and high-stage adenocarcinomas demonstrate enhanced expression of CD55 and CD59, indicating a poor prognosis. It is essential for pathologists to meticulously perform grossing and reporting of CRC and dispatch the histopathology report after proper clinicopathological correlations. These immunomarkers can also be included in a routine IHC panel for prognostic and therapeutic purposes.

Keywords

Adenocarcinoma, Immunohistochemistry, Membrane bound complement regulatory proteins, Tumour grade, Tumour stage

Cancer is undoubtedly a worrisome disease in our country, leading to increased morbidity and mortality worldwide. The main cause of mortality associated with this cancer is tumour invasion and metastasis. There are many factors that determine the prognosis of this cancer, with two important factors being the level of invasion and lymph node metastasis. Node-negative disease usually shows a favourable clinical course. It is also observed that approximately one-third of node-negative CRC cases show recurrence and disease progression. It is probably due to a failure to detect occult disease (1). During the progression of the tumour, aberrant genetic changes occur early. To predict the potentiality of tumour metastasis, molecular profiling of specific tumour markers in the primary tumour may be helpful (2). There are different complement proteins in our body that help fight against tumour formation. Among them, some are known as Decay Accelerating Factor (DAF) or membrane-bound Complement Regulatory Proteins (mCRPs), for example, CD55, CD59, and CD46. Their roles are currently being studied by immunohistochemistry. Recent studies report that aberrantly expressed mCRPs in cancer cells escape from complement-mediated cell lysis (1),(2),(3). The aberrantly expressed or upregulated mCRPs in some cancer cells inhibit the activation of complements and the formation of the Membrane Attack Complex (MAC). They also block the activation of T cells, their proliferation, and differentiation. In this way, they help tumour cells escape from immune attack, leading to poor prognoses for patients with tumour (4).

It has been demonstrated that novel anti-CD55 and CD59 antibodies suppress colorectal tumorigenesis in subsets of colorectal cancer tissues. These antibodies activate different complements and cause cancer cell death through the production of proinflammatory cytokines (5). Additionally, they can kill cancer cells by expanding natural killer cells and/or stimulating macrophage-mediated antibody-dependent cell-mediated cytotoxicity. The discovery of 5-fluorouracil (5-FU) in 1957 was a landmark advance for CRC patients and has since been used as one of the first-line treatments in advanced CRC (6). However, many cases are facing resistance to it (5). Importantly, combinational therapy with these novel anti-CD55 and anti-CD59 antibodies with 5-FU may enhance the therapeutic effect against CRC. Interestingly, the stools of CRC patients contain soluble CD55 and CD59, possibly mediated by metalloproteinase-7 (6). This suggests that these immunomarkers may be used as a markers for CRC, and these antibodies could potentially be utilised for CRC diagnosis and therapeutics.

Therefore, the primary aim was to assess the expression of CD55 and CD59 in colorectal cancer, while objectives were:

1. To determine the clinical parameters of the patients.
2. To identify the histological findings of different CRC
3. To determine the grading and staging (TNM) of CRC.
4. To evaluate the expression of CD55 and CD59 in histologically diagnosed carcinomatous colorectal tissue.
5. To examine the association of CD55 and CD59 expression with different clinical parameters (age, sex, tumour location, and tumour type) as well as the grade and stage of CRC.

Material and Methods

The present study was a prospective, cross-sectional, institution-based study conducted in the Department of Pathology in collaboration with the Department of General Surgery at Nil Ratan Sircar Medical College and Hospital (NRSMCH), Kolkata, West Bengal, India, for a period of 18 months (February 2020 to July 2021). The total number of study participants in this study was 90. The study was approved by the Institutional Ethical Committee (IEC) and was assigned the IEC number: Memo number- NO/NMC/693, Dated-10.02.2020.

Inclusion criteria: All specimens of colorectal growth obtained from all types of colectomy, abdominoperineal resection, and anterior resection, which were subsequently diagnosed histologically as CRC were included in the study.

Exclusion criteria: Benign and inflammatory (non malignant) lesions of the colon, malignant lesions not arising from the colon were excluded from the study.

Study Procedure

While undertaking this study, voluntary written informed consent was obtained from each patient, and full confidentiality of the patients was maintained. The study included all admitted indoor patients with a preoperative diagnosis of adenocarcinoma by colonoscopic biopsy. All types of colectomies, abdominoperineal resection, and anterior resection were performed in the Department of General Surgery at NRSMCH, and corresponding specimens were received in the Department of Pathology, NRSMCH, meeting the inclusion criteria during the study period. The grossing and reporting of the specimens were conducted according to the College of American Pathologists (CAP) Protocol (1).

The biopsy procedure involved removing the affected colorectal tissue from the patients under anaesthesia and sending it to the laboratory in an adequate amount of 10% Neutral Buffer Formalin (NBF) (1:10 dilution of a 37% formaldehyde stock solution) with proper labeling. The samples were kept for proper fixation for a minimum of 24 hours followed by grossing. Representative sections were taken from the tumour proper (3 sections-full thickness, including serosa), adjacent apparently uninvolved colonic mucosa (1 section), resection margins (proximal, distal, circumferential, if applicable), omental tissue (3 sections to look for any tumour deposit), and lymph nodes in the attached mesentery. Characteristics such as colour, consistency, necrosis, haemorrhage, calcification, etc., were also noted. A meticulous search was conducted for lymph nodes sent separately. After grossing, the tissue was dehydrated in graded ethyl alcohol (100%, 90%, 80%, 70%), cleared with xylene, and embedded in liquid paraffin (melting point 50-60°C) for proper block preparation in a Leuckart mold block using paraffin. sections were cut at a thickness of 4 μm using a Leica rotary microtome followed by Haematoxylin and Eosin (H&E) staining. Broders grading and staging (TNM) of CRC were performed through histopathological examination (1). Grade-1 indicates a tumour with >95% gland formation, grade-2 indicates 50-95% gland formation, and grade-3 indicates 0-50% gland formation (1).

Immunohistochemical methods and analysis (1): For the immunohistochemical analysis of CD55 and CD59, 3.0 μm paraffin sections were taken on poly-L-lysine-coated slides and deparaffinised in xylene, followed by hydration in descending grades of ethanol. Antigen retrieval was performed by heating sections at 95°C (3 cycles of 5 minutes each for CD55 and CD59) in citrate buffer using an antigen retriever system (BioGenex, USA). Sections were then incubated with power block (Biogenex, USA) for 10 minutes to reduce non specific antibody binding, followed by incubation with primary antibodies (CD55-Rabbit polyclonal antibody and CD59-Rabbit polyclonal antibody) for one hour at 4°C. After three washes with Trisphosphate Buffer Solution (TBS), a secondary antibody was added and incubated for 30 minutes. After a further three washes with TBS, 3,3´-diaminobenzidine substrate (DAB tetrahydrochloride) was applied to the sections for 10 minutes, and the sections were counterstained with Haematoxylin, dehydrated with ethanol and Xylene, and permanently mounted with DPX. In all the above immunostaining, cell counting was done in the areas where the tumour was most prominent. Areas of haemorrhage and necrosis were avoided. The immunohistochemical reports were mostly completed within three weeks after the clinical correlation of each case.

Internal control: Normal colonic mucosa.

Positive control: Previously known case of CRC (CD55 and CD59 positive case).

Negative control: Omitting the primary antibody.

The expression of CD55 and CD59 was observed at the membrane and cytoplasm.

A standard immunoreactivity scoring system was followed (7), based on: a) the percentage of stained cells (0 points-4 points); and b) the intensity of stained cells (0 points-3 points).

a) Percentage of stained cells (0 point- 4 points).

0 5% or less
1 6% to 25%
2 26 % to 50%
3 51% to 75%
4 Greater than 75%

b) Intensity of stained cells (0 point-3 points).

0 No stain
1 Mild (buff)
2 Moderate (yellow)
3 Strong (brown)

a and b were multiplied together to obtain the final score. The staining score was stratified as weak (score range 0 to 4) or strong (score range 5 to 12) based on the proportion and intensity of positively stained cancer cells.

The following parameters were evaluated for association of immunomarkers (CD55 and CD59:

1) Clinical parameters (age, sex, clinical presentations, their duration, food habits, family history of CRC, prior chemotherapy, etc.).
2) Histopathological findings (type of colorectal cancer, their differentiation/grading, and staging) were evaluated.

Statistical Analysis

For statistical analysis, data were entered into Microsoft Excel (MS). The data were summarised as numbers, means±standard deviations (SD) for numerical variables and as percentages (%), and ranges for categorical variables. The Chi-square test was used to determine the significance of the study. Data analysis was conducted using SPSS version 27.0 (IBM, Illinois, US) and GraphPad Prism version 5. A p-value of ≤0.05 was considered statistically significant.

Results

A total of 90 CRC patients were included in this study, with the maximum number of patients, 30 (33.30%), belonging to the 51-60 age group (Table/Fig 1). The mean age was 52.32±14.007 years, with the minimum and maximum ages being 16 years and 79 years, respectively.

Male preponderance was observed, with 56 (62.2%) cases being male. The number of male and female patients were 56 (62.2%) and 34 (37.8%), respectively (Table/Fig 2).

Symptoms are shown in (Table/Fig 3). Only 09 (10.%) patients had a positive family history of CRC. A total of 77 (85.6%) patients consumed a non vegetarian diet, while 13 (14.4%) patients consumed a vegetarian diet.

The rectum was the predominant site of involvement in 32 (34.4%) patients, while the descending colon (01) was the least involved site with only 01 (01.11%) patient (Table/Fig 4).

In terms of histological types, 77 (85.6%) patients were diagnosed with adenocarcinoma-No Special Type (NST), other types are shown in (Table/Fig 5).

Among the adenocarcinomas, distribution of well, moderate and poorly differentiated is shown in (Table/Fig 6),(Table/Fig 7),(Table/Fig 8),(Table/Fig 9).

The majority of patients, 44 (48.9%), were in stage three (Table/Fig 10). A total of 36 (40.00%) patients had stage pT3N0M0, followed by 26 (28.88%) with pT3N1M0, 05 (05.6%) with pT2N0M0, 04 (04.4%) with pT3N1bM0, pT3N1aM0, and pT4aN0M0 each, 03 (03.3%) with pT1N1M0, pT2N1aM0, and pT3N2aM0 each, and 02 (02.2%) with pT4N0M0.

Strong and weak intensity of CD55 and CD59 is shown in (Table/Fig 11).

Regarding the expression of CD55 with the grade of the tumour, mostly weak expression was observed in well-differentiated adenocarcinomas.

Among the 59 moderately differentiated carcinomas, 42 (71.18%) exhibited strong expression, and 17 (28.82%) showed weak expression. All 18 poorly differentiated carcinomas showed strong expression (100.00%) (Table/Fig 12).

Concerning the expression of CD59 with the grade of the tumour, predominantly weak expression of CD59 was noted in well-differentiated adenocarcinomas. Out of 59 cases of moderately differentiated adenocarcinomas, 46 cases (77.97%) showed strong expression, while 13 cases (22.03%) exhibited weak expression. Among the 18 poorly differentiated adenocarcinoma cases, 17 cases (94.44%) demonstrated strong expression

Regarding the expression of CD55 with the stage of the tumour, Stage-III and Stage-IV cancers exhibited stronger expression of CD55 (44 cases, 91.66% out of 48 cases) compared to Stage-I and Stage-II cancers (18 cases, 42.85% out of 42 cases) (Table/Fig 14).

In this study, Stage-III and Stage-IV cancers showed stronger expression of CD59 (46 cases, 95.83% out of 48 cases) than Stage-I and Stage-II cancers (19 cases, 45.23% out of 42 cases) (Table/Fig 15). The association of both marker expressions with TNM staging and histological grade was found to be statistically significant (Table/Fig 16).

Discussion

The CRCs are the third most common carcinoma worldwide (8). As a result, many studies and research works have been done and are being conducted. More frequently, some newer parameters and markers are being used to detect and prognosticate these tumours as our knowledge regarding CRCs is continuously updated. Enhanced CD55 expression is found not only in CRC (9) but also in malignancies of the lung (10), stomach (11), breast (12), ovary (13), prostate (14), cervix (15), and leukemia (16). High expression of CD55 and CD59 is a poor indicator of colorectal cancer (17),(18). One should also keep in mind that CD55 is also expressed in malaria, protein-losing enteropathy, and some autoimmune diseases (19),(20),(21).

Cancer is strongly age-related. The incidence rises progressively with age. Present study showed that the mean±SD age of the study population was 52.32±14.007 years. The majority belonged to the 50 to 60 years age group with a minimum age of 16 years and a maximum age of 79 years. Out of 90 cases, 18 (20.00%) cases were from the age group <40 years, while the remaining 72 (80.00%) cases were above 40 years of age similar to the study by Patil SP et al., (22). In their study, the mean age of the patients was 47.20 years, and the age range was 11 years to 85 years. The mean age at diagnosis was 58.4 years, and the range was 23-85 years in the study of Peedikayil MC et al., (23).

The present study showed a slight increase in the incidence of CRC in males, 56 (62.2%), compared to females, 34 (37.8%), resulting in a male-to-female ratio of 1.64:1. Similar results were reported by Nayak SP et al., (Male-40, female-26, M: F ratio-1.53:1) (24), and Laishram RS et al., (Male-50, female-31, M: F ratio-1.61:1) (25).

In CRC, dietary factors play an important role; there is a close correlation between meat consumption and the incidence of large-bowel cancer. In this study, 13 patients (14.4%) were vegetarian, and the remaining 77 cases (85.6%) were non vegetarian, similar to the study by Laishram RS et al., (vegetarian-21, 15.75% and non-vegetarian-37, 84.25%) (25). A positive family history was noted in only 09 cases (10.00%), similar to the study by Nayak SP et al., (20 cases, 14.56%) (24). Symptomatically, 44 (48.9%) patients presented with rectal bleeding, 28 (31.1%) with abdominal pain, and 18 (20.00%) with altered bowel habits, similar to the findings of the study by Patil SP et al., (n=62, rectal bleeding 28 (45.16%), abdominal pain 20 (32.25%), and altered bowel habits in 10 cases (16.12%) (22). The most common site of the lesion was the rectum 32 (35.55%), followed by the caecum 27 (30.00%), sigmoid colon 15 (16.66%), ascending colon 11 (12.22%), transverse colon 04 (04.44%), with the rarest being the descending colon 01 (01.11%) (Table/Fig 4). Similar findings were observed in other studies as well (22),(25),(26),(27),(28). Peedikayil MC et al., found that most of the tumours (N= 163, 74.00%) were located distal to the splenic flexure (23). Nayak SP et al., found the commonest site of involvement to be the sigmoid colon {N=45, 25 (37.00%)}, followed by the caecum (10) and rectum (10) 18.50% each (24).

Microscopically, adenocarcinoma- NST was the most common histological type, accounting for 77 (85.6%) cases, followed by mucinous adenocarcinoma 12 (13.6%) and signet ring cell carcinoma 1 (01.1%) in the present study. Among the adenocarcinoma NST category, moderately differentiated tumours were the most frequent, with 59 (65.6%) cases, followed by poorly differentiated with 18 (20%) and well-differentiated tumours with 13 (14.4%) cases (Table/Fig 6). This finding was similar to the study by Wong SCC et al., (29). In the present study, the most common TNM staging was Stage-III, followed by Stage-II (Table/Fig 10). This was in accordance with the observations of Wong SCC et al., {Stage-III-48 (52.76%), Stage-II-32 (42.56%)} (29) and Jass JR {Stage-III- 41 (42.76%), Stage-II-35 (40.96%)}, respectively (30). Mesenteric lymph nodes were tested positive in 47 (52.22%) cases. The maximum number of lymph node involvements in this study was 09 in a single case. Out of 90 cases, only 05 (05.55%) patients had distant metastasis, with 03 patients having metastasis in the liver, 01 patient in the lung, and 01 female patient in the brain.

Immunohistochemically, CD55 was abundant in colorectal cancer tissue, but its intensity of expression was much lower in normal colonic tissue. Out of 90 histologically proven normal tissues, weak expression of CD55 was noted in 82 tissues (91.11%) and strong expression in 8 tissues (08.89%), while among 90 histologically proven cancerous tissues, 62 (68.9%) had strong expression and 28 (31.1%) had weak expression of CD55 (Table/Fig 11). This result was similar to the study by Koretz K et al., {n=88, 57 (64.77%) strong and 31 (35.23%) weak} and Shang Y et al., {n=56, 43 (76.78%) strong and 13 (23.22%) weak} [9,31]. In this study, the association of the expression of CD55 with the age of the patient (p-value=0.1097), sex of the patient (p-value=0.7861), location of the tumour (p-value=0.7371), and histological type of the tumour (p-value=0.0769) were not significant. These findings were in concordance with the findings of the study done by Shang Y et al., (p-value=0.1207, p-value=0.5462, p-value=0.6545, p-value=0.0834) (31). The association of the grade of the tumour with the expression of CD55 was statistically significant. The chi-square value and p-value were 25.6386 and 0.0001, respectively (Table/Fig 15). These findings were consistent with the study conducted by Shang Y et al., (p-value=0.0054) (31). While CD55 upregulation in CRC has been demonstrated in some studies [9,32,33], several groups have not found high expression of CD55 in colorectal tumour tissues (34). Regarding the expression of CD55 with the stage of the tumour, a statistically significant association was found (p-value=0.0013) with a chi-square value of 25.3372 (Table/Fig 15), similar to the study by Shang Y et al., (p-value=0.0034) (31).

Immunohistochemically, the expression of CD59 was abundant in CRC tissue, but its intensity of expression was much lower in normal colon tissue. Out of 90 histologically proven normal tissues, weak expression of CD59 was found in 84 tissues (93.3%) and strong expression in 06 tissues (06.7%), while among 90 histologically proven cancerous tissues, 65 cases (72.22%) had strong expression and 25 cases (27.78%) had weak expression of CD59 (Table/Fig 11), similar to the study by Shang Y et al., {N-56, 41 (73.21%) strong, 15 weak (26.79%)} (31).

No significant association of the expression of CD59 with the age of the patient (p-value=0.5179), sex of the patient (p-value=0.0945), location of the tumour (p-value=0.0582), and histological type of the tumour (p-value=0.1566) was noted. These findings were consistent with the study conducted by Shang Y et al., (p-value=0.6712, p-value=0.1013, p-value=0.0654, p-value=0.1656) (31). A significant association of the expression of CD59 with the grade of the tumour was found in this study. The association of the grade of the tumour with the expression of CD59 was statistically significant (p-value=0.0001) with a chi-square value of 26.3348 (Table/Fig 15), similar to the study conducted by Shang Y et al., (p-value=0.0002) (31). In this study, Stage-III and Stage-IV cancer exhibited stronger expression than Stage-I and Stage-II cancer. A statistically significant association between the stage of the tumour and the expression of CD59 (p-value=0.0001) with a chi-square value of 31.4685 was found (Table/Fig 15), similar to the study by Shang Y et al., (p-value=0.0003) (31). Watson NF et al., observed the expression of CD59 in 69 (15.00%) of cases overall, and it was significantly associated with tumour grade but not with tumour site, stage, or histological type (18). Fonsatti E et al., had given major emphasis on CD59 extensively on solid malignancies, and clinical approaches of humoral immunotherapy also had been implemented by them (35).

Limitation(s)

Survival analysis of the patients (to understand the prognosis) as well as the follow-up of the patients was not conducted. The study did not focus on the association of CD46 (a type of DAF) with the parameters mentioned above. The stool of CRC patients was not examined for soluble CD55 due to limited resources in this study. All cancer patients were treated with 5-FU and not with novel anti-CD55 or CD59 antibody drugs. Since this study was conducted at a single centre in Kolkata, the results cannot be generalised to all regions.

Conclusion

The incidence of CRC is gradually increasing in our country. Since the clinical and histopathological presentations vary from case to case, clinicopathological association is crucial for prognostic purposes. Immunohistochemical associations of CD55 and CD59 can be performed in CRC, as high-grade (poorly differentiated) and high-staged adenocarcinoma commonly show increased expression of these new markers, ultimately leading to a poor prognosis for the patient. This may help in guiding the early initiation of treatment in the future.

Acknowledgement

Authors are indebted to Mr. Animesh Hazra (Assistant Professor, Computer Science and Engineering, Jalpaiguri Govt. Eng. College, West Bengal), Prof. Dr. Nibaran Das (Computer Science and Engineering, Jadavpur University, West Bengal) and Mr. Soumyajyoti Dey (PhD Scholar, Computer Science and Engineering, Jadavpur University, West Bengal) and Mr. Ranit Hazra (BA, Geography, Honours, Ashutosh College, Kolkata, West Bengal) for their technical support.

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DOI and Others

DOI: 10.7860/JCDR/2024/69311.19379

Date of Submission: Dec 28, 2023
Date of Peer Review: Jan 31, 2024
Date of Acceptance: Mar 05, 2024
Date of Publishing: May 01, 2024

AUTHOR DECLARATION:
• Financial or Other Competing Interests: None
• Was Ethics Committee Approval obtained for this study? Yes
• Was informed consent obtained from the subjects involved in the study? Yes
• For any images presented appropriate consent has been obtained from the subjects. Yes

PLAGIARISM CHECKING METHODS:
• Plagiarism X-checker: Dec 28, 2023
• Manual Googling: Feb 26, 2024
• iThenticate Software: Mar 01, 2024 (10%)

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EMENDATIONS: 7

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