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.
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 : 2022 | Month : August | Volume : 16 | Issue : 8 | Page : EC47 - EC51 Full Version

Association between Preoperative RBC Parameters with Serum VEGF in Women Diagnosed with Breast Carcinoma: A Case-control Study


Published: August 1, 2022 | DOI: https://doi.org/10.7860/JCDR/2022/57026.16806
Apoorva P Gowda, TS Rekha, MVSST Subbarao, Venugopal R Bovilla

1. Postgraduate Student, Department of Pathology, JSS Medical College, JSS Academy of Higher Education and Research, Mysuru, Karnataka, India. 2. Associate Professor, Department of Pathology, JSS Medical College, JSS Academy of Higher Education and Research, Mysuru, Karnataka, India. 3. Professor, Department of Biochemistry, Center of Excellence in Molecular Biology and Regenerative Medicine, JSS Medical College, JSS University, Mysuru, Karnataka, India. 4. GHES LMIC Fellow, Department of Global Health Equity Scholars Program, School of Public Health, UC Berkeley, USA.

Correspondence Address :
Dr. TS Rekha,
Associate Professor, Department of Pathology, JSS Medical College, SS Nagar, Mysuru-570015, Karnataka, India.
E-mail: rekhats12@gmail.com

Abstract

Introduction: The complete blood count is the first investigation to be done in every patient with breast carcinoma before surgery. Vascular Endothelial Growth Factor (VEGF) plays a major role in angiogenesis, metastasis and progression of tumours.

Aim: To assess Red Blood Cell (RBC) parameters in breast carcinoma patients and controls, and to evaluate its relation with serum VEGF.

Materials and Methods: The present study was a case-control study, conducted in Department of Pathology of JSS Medical College and Hospital, Mysuru, Karnataka, India from November 2019 to April 2021. Preoperative venous blood samples were collected and run in an automated analyser Mindray CAL-6000 for all haematological parameters. Preoperative serum samples were collected and serum VEGF was estimated using the Enzyme Linked Immunosorbent Assay (ELISA) method. Statistical analysis was performed with Statistical Package for the Social Sciences (SPSS) version 22.0 to evaluate the association between RBC parameters and serum VEGF using Mann-Whitney U test.

Results: A total of 80 samples were evaluated, which included 40 preoperatively diagnosed breast cancer cases and 40 age and sex matched controls. RBC parameters such as RBC, Haemoglobin (Hb), Haematocrit (Hct), red cell indices Mean Corpuscular Volume (MCV), Mean Corpuscular Haemoglobin (MCH), Mean Corpuscular Haemoglobin Concentration (MCHC), reticulocytes, Red cell Distribution Width (RDW) and nucleated RBCs (nRBCs) were measured. The nRBCs and serum VEGF of cases were significantly higher than controls. A statistically significant association between patients with low Hb and high serum VEGF was found.

Conclusion: A higher percentage of breast carcinoma cases with anaemia in the present study was significantly associated with high serum VEGF, reflecting anaemia induced hypoxia may trigger the tumour cells to secrete VEGF.

Keywords

Anaemia, Haemoglobin, Hypoxia, Red blood cell, Vascular endothelial growth factor

Amongst the simple routine tests performed prior to breast cancer surgery, haematology profile is one of the important tests. RBC parameters which include RBCs, Hb, Hct, red cell indices, reticulocytes, RDW and nRBCs are measured. The analysis of RBC parameters is used to detect anaemia, type of anaemia, response of the body to anaemia and breast cancer. Angiogenesis is the process of forming new blood vessels from an existing vascular network in response to hypoxia, which is required for tumour growth, invasion, and spread (1). The most effective endothelial cell mitogen and a regulator of vascular permeability ie., VEGF, induces angiogenesis and has emerged as a potent tool in the prognosis (2). Patients with initial-stage of breast tumours with high levels of VEGF have a higher risk of relapse or mortality than patients with tumours that are less angiogenic (3).

It is known that in anaemia, there is decrease in Hb and RBC leading to hypoxia especially in the growing tumour cells. It has been observed that, in cancer patient Hb and Hct are linked to a greater risk of heart failure (4). But it is not clearly known whether there is a relationship between the RBC parameters and serum VEGF levels in breast carcinoma. The novelty of present study was to know whether alteration in RBC parameters that results in hypoxia can affect the serum VEGF levels which may alter the prognosis of breast cancer patients.

The objective of the present study was, the analysis of RBC parameters in breast cancer patients and healthy controls, comparing with serum VEGF in breast cancer to elucidate any association between them.

Material and Methods

A case-control study was conducted from November 2019 till April 2021 in the Department of Pathology of JSS Medical College and Hospital, Mysuru, Karnataka, India. All procedures performed were in accordance with the ethical standards of the Institutional Research Committee (JSS/MC/PG/5189/2019-20) and the 1964 Helsinki declaration. Ethical approval was obtained by the Institutional Ethical Committee (IEC) of JSS Medial College and Hospital, Mysuru, Karnataka, India.

Inclusion criteria: Forty preoperatively diagnosed breast carcinoma patients either by fine needle aspiration or core needle biopsy were collected and taken as cases. Forty healthy and age matched female volunteers attending general health checkup (without history of breast carcinoma) were included as controls.

Exclusion criteria: Women with postoperative diagnosis of breast carcinoma (preoperative diagnostic test done in other Institute), serum sample collected but underwent operation in other hospital, benign breast disease, postchemotherapy cases were excluded from the study.

Venous sample in EDTA vacutainers and serum samples were collected after informed consent under aseptic precautions. EDTA samples were run in automated analyser Mindray CAL-6000 and the data was collected for RBC parameters such as RBC, Hb, Hct, MCV, MCH, MCHC, reticulocytes, RDW and nRBCs. Serum was separated by centrifuging the blood for 10 minutes at 1500 rpm, and stored at -80o C in a deep freezer. RayBio® Human VEGF-A Enzyme Linked Immunosorbent Assay (ELISA) Kit was used to estimate serum VEGF by sandwich ELISA method.

Statistical Analysis

Statistical analyses were performed using SPSS 22.0 statistical package for Windows. Continuous data were expressed as mean, median, standard deviation and range, while categorical data were presented as numbers and percentages. The distribution of variables was checked with the Kolmogorov Smirnov test. The categorical parameters between cases and controls were compared with Mann-Whitney U test. Receiver Operating Characteristic (ROC) curve analysis was performed to determine the cut-off of serum VEGF to divide the cases with low and high VEGF levels. The p-values <0.05 was considered statistically significant.

Results

Age distribution in cases ranged from 34 to 85 years with a mean±age of 52.33±12 years. Age distribution in controls ranged from 30 to 74 years with a mean age of 52±12 years with no significant differences between cases and controls (p=0.92).

On comparing the RBC counts, both cases and controls had low and high RBC counts (Table/Fig 1). However, percentage of cases with low RBC counts (17.5%; 7/40) were higher than controls (10%; 4/40). There were equal number of cases and controls with high RBC counts. Mean RBC count was slightly lower among cases (4.2±0.5×106/uL) than controls (4.3±0.5×106/uL), which was not a statistically significant (p=0.689). There was no statistically significant difference in the mean Hb level between cases (11.6±1.6 g/dL) and controls (11.9±1.5 g/dL) (p=0.391). But the percentage of cases (50%;20/40) with low Hb (<12g/dL) was slightly higher than controls (47.5%; 19/40) (Table/Fig 1). In cases, 70% (28/40) of them had Hct in normal range and 30% (12/40) had low Hct; whereas among controls 52.5% (21/40) had normal Hct and 47.5% (19/40) had low Hct. Mean Hct had no statistically significant difference (p=0.452) (Table/Fig 1).

The majority of cases and controls had MCV, MCH and MCHC within normal ranges and their mean values had no significant difference (Table/Fig 1). The percentage of cases with low MCV and MCH were 7.5% (3/40) and 15% (6/40) respectively. Similarly, the percentage of controls with low MCV and MCH were 30% (12/40) and 40% (10/40) respectively. Equal percentage of cases and controls had high MCH. All cases and controls had MCHC within normal range (18-48 g/L). The mean MCHC of cases were slightly higher (33±1.3 g/L) than controls (31.8±4.7 g/L), however there was no statistically significant difference (p=0.105).

In 77.5% (31/40) of cases and 57.5% (23/40) of controls had RDW in normal range (Table/Fig 1). Mean RDW was low in cases (13.8±2.0) than controls (14.4±1.9) but the difference was statistically not significant (p=0.211). Mean reticulocyte counts were within normal ranges in both cases and control with no significant differences (Table/Fig 1). The mean nRBCs were significantly high in cases (0.19±0.1) than controls (0.02±0.01) with a p-value of 0.008.

Mean serum VEGF of cases (100.7±33.91 pg/mL) were significantly higher than control (56.4±9.77 pg/mL) with a p-value of 0.013. Median serum VEGF of cases were significantly higher-70.4 (145.5-103.8) pg/mL than controls-44.3 (37.4-83.8) pg/mL with a p-value of 0.007. On statistical analysis, it was found that the VEGF values were not normally distributed hence median value was taken into consideration.

ROC analysis was done and area under the curve was 0.674, serum VEGF of 72.3 pg/mL was taken as cut-off value with a sensitivity of 47.5% and specificity of 70% with p-value of 0.007 (Table/Fig 2). Considering 72.3 as cut-off value, cases were categorised into low VEGF and high VEGF categories respectively and were compared with RBC parameters.

The mean serum VEGF in patients with low VEGF category was 29.4, 48.7 and 49.1 pg/mL when RBC counts were low, normal and high respectively. Similarly, the mean serum VEGF among high 48VEGF category was 83.6, 119.8 and 99.6 pg/mL, respectively. Absence of association between low, high and normal RBC counts and serum VEGF categories were observed (Table/Fig 3)a,(Table/Fig 4).

Among patients with Hb <12, the mean serum VEGF was 48.5 and 95.1 pg/mL in low and high VEGF category respectively. A statistically significant association between patients with low Hb and high serum VEGF with a p-value of 0.015 was found. Among patients with normal Hb, the mean serum VEGF was 45 and 126.6 pg/mL in low and high VEGF category respectively and observed no association between these categories (Table/Fig 3)b,(Table/Fig 4).

Among patients with Hct<35, the mean serum VEGF was 29.7 and 80.0 pg/mL low and high VEGF category respectively. Similarly, when Hct was in normal range, the mean serum VEGF was 39.1 and 78.4 pg/mL, respectively. There was no association between low and normal Hct with serum VEGF categories (Table/Fig 3)c,(Table/Fig 4).

The mean serum VEGF, among patients in low and high VEGF categories was compared with different categories of MCV, MCH and MCHC without statistically significant association (Table/Fig 3)d,(Table/Fig 3)f,(Table/Fig 4). In breast carcinoma cases with normal RDW, the mean serum VEGF in low and high VEGF categories were 47 and 99.6 pg/mL respectively. Likewise, when the RDW was high the mean serum VEGF in low and high VEGF categories were 48.7 and 334.6 pg/mL respectively. On statistical analysis, there was no association between any of these categories (Table/Fig 3)g,(Table/Fig 4).

All the cases in this study had normal reticulocyte counts: the means serum VEGF levels were 55.6 and 82.1 pg/mL in low and high VEGF category, respectively (Table/Fig 3)h,(Table/Fig 4). Amongst patients with nRBCs, significantthe mean serum VEGF were 60.5 and 88.3 pg/mL in low and high VEGF category respectively (Table/Fig 3)i,(Table/Fig 4). Association between either reticulocyte count or nRBCs with serum VEGF levels, were not noticed.

Discussion

Breast cancer is the most common invasive malignancy and the second leading cause of tumour-related deaths among women globally (5). RBC parameters are one of the initial tests requested for all patients with breast cancer to detect anaemia and its associated changes. In this study, RBC parameters in both breast carcinoma cases and healthy controls were assessed. These were then compared with a potential breast cancer progression risk indicator-serum VEGF (6). Growth of tumour requires supply of many new blood vessels in order to provide and support the metabolic activity; these growing tumours secrete many growth factors, including VEGF to recruit new blood vessels (7). This stimulates proliferation of endothelial cells and formation of new vessels called angiogenesis (8),(9). VEGF receptor system is a primary tumour angiogenesis regulator and it has been resulted in progression and metastasis of the tumour (10),(11). Inhibition of VEGF results in suppression of development and metastasis of tumour (12).

In the present study, serum VEGF by sandwich ELISA method was estimated and found to be significantly elevated in cases than controls. Present study results were consistent with Lawicki S et al., and Ali EM et al., findings (13),(14). The preoperative detection of serum VEGF may support in identifying the severity and prognosis of breast cancer cases (12). Statistically, 72.3 pg/mL as a cut-off for serum VEGFwas derived and thereby all breast carcinoma cases were divided into low VEGF and high VEGF categories. In the current study, similar mean age in cases and controls was found. Mean age of present study was comparable with Okuturlar Y et al., Raghunathachar Sahana K et al., and Harano K et al., (15),(16),(17).

Low RBC count and Hb, the indicators of anaemia were observed in both cases and controls. However, the high percentage of cases had low RBC count and low Hb than those of controls. This indicates that the anaemia which is prevalent in women in developing country like India (18) probably due to lower socio-economic status and poor nutrition was aggravated in breast carcinoma patients (19). Metastasis to the bone marrow from breast cancer and nutritional deficiency induced by anorexia can be associated with suppression of erythropoiesis resulting in anaemia (4),(20). While in the present study Hb<12 was in 50% (20/40) of cases, Rana APS et al., has reported 60% and Zhang Y et al., in 25.2% (4),(21). High RBC count among controls could be due to dehydration or on anaemia treatment. It has been reported that some of the breast carcinoma tumour cells secrete erythropoietin that can result in high RBC and Hb values (22).

In the present study, association between RBC counts and serum VEGF categories was not noticed. Surprisingly the serum VEGF was low in patients with low RBC count than patients with normal RBC count. In contrast Bhatta SS et al., have reported inhibition of serum VEGF with antiangiogenic therapy increased erythropoiesis and increased RBCs (23). Nevertheless, in patients with low Hb, a significant association with high serum VEGF was found, upholding the previous reports that low Hb results in hypoxia which in turn stimulates secretion of VEGF (22). Majority (97%) of the oxygen supply to tissues are derived from oxygen bound to haemoglobin. Therefore, in anaemia cases, low haemoglobin levels might increase hypoxia in tumours (22). Zhang Y et al., have reported that preoperative anaemia in breast carcinoma patients was associated with poor prognosis (21).

Another method of detecting anaemia was by estimation of Hct, which is the percentage of RBCs in total volume of blood. In contrast to Hb and RBC count, the percentage of patients with low Hct was less compared to controls in the present study. It has been reported that Hb was a more accurate method than Hct in assessing anaemia, which could be the cause of contrasting results (24). The mean Hct in cases were higher than in controls and similar values were published by Rana APS et al., (4). Statistical analysis revealed lack of association between Hct and serum VEGF.

Erythrocyte indices such as MCV, MCH and MCHC provide a clue to the different types of anaemia (25). Mean erythrocyte indices were within normal range in this study and a similar observation was noted in previous study (4). Erythrocyte indices showed that majority of the cases had normocytic normochromic anaemia while in controls microcytic hypochromic anaemia was more prevalent in this study. Previous study has reported that one of the causes for normocytic normochromic anaemia was breast cancer (26) which substantiates the reason for majority of cases with normocytic normochromic anaemia. The findings in the present study that mean erythrocyte indices of cases were higher than the controls contrasted with the findings of Akinbami A et al., (27). Low serum VEGF levels was detected in patients with decreased MCV than those with normal MCV range. Those patients with decreased MCH had high VEGF than those with normal and high MCH. However, association between erythrocyte indices and serum VEGF was not found.

The RDW is a parameter to measure the variability of the size of the circulating RBCs (25). Both the percentage and mean RDW were lower in cases compared to controls indicating that the RBCs in cases were more homogenous than controls in the current study. However, Seretis et al., reported higher RDW in cases than controls (28). The presence of varied types of RBCs such as microcytes, normocytes and reticulocytes found in controls resulted in high RDW. The reticulocytes and nRBCs representing the immature erythrocytes are either low or not normally found in peripheral blood respectively. Even though there was no increase in reticulocytes in the present study, nRBCs were significantly increased in cases than controls. Increased reticulocytes and presence of nRBCs indicates increased erythropoiesis in bone marrow found in patients as a response to anaemia (29). In peripheral blood, presence of nRBCs has shown to be associated with various medical conditions like solid cancers, haematological malignancies, cardiovascular conditions, infections, haemorrhage etc., (30),(31). An association of serum VEGF with RDW, reticulocytes or nRBCs was not observed in this study. On reviewing the literature, studies analysing the association between all the RBC parameters (except for Hb) and serum VEGF were not found to the best of the author’s knowledge.

Limitation(s)

The limitation of the present study includes small sample size and lack of demonstration of VEGF induced angiogenesis in breast carcinoma patients with anaemia. Therefore, it is recommended to conduct similar studies with large sample size at various geographic locations and further investigate the extent of angiogenesis in breast carcinoma patients with and without anaemia.

Conclusion

Anaemia is prevalent in elderly women due to various causes in developing country like India, which can be aggravated by breast carcinoma. Either as a compensatory mechanism to anaemia leading to increased bone marrow erythropoiesis or irritation due to metastasis into bone marrow had significant increase in nRBCs in the peripheral blood of breast carcinoma patients. A higher percentage of breast carcinoma cases with anaemia in the present study was significantly associated with high serum VEGF, reflects anaemia induced hypoxia may trigger the tumour cells to secrete VEGF.

Acknowledgement

Authors would like to acknowledge (a) the infrastructure support provided by Department of Science and Technology to CEMR Laboratory (CR-FST-LS-1/2018/178) and to Department of Biochemistry (SR/FST/LS-1-539/2012); (b) the laboratory facilities provided by CEMR laboratory (DST-FIST supported centre), Department of Biochemistry (DST-FIST supported department), and Special Interest Group on Cancer Biology and Cancer Stem Cells (SIG-CBCSC), JSS Academy of Higher Education and Research (Mysore, Karnataka, India).

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DOI and Others

DOI: 10.7860/JCDR/2022/57026.16806

Date of Submission: Apr 11, 2022
Date of Peer Review: Apr 23, 2022
Date of Acceptance: Jul 14, 2022
Date of Publishing: Aug 01, 2022

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. NA

PLAGIARISM CHECKING METHODS:
• Plagiarism X-checker: Apr 16, 2022
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• iThenticate Software: Jul 09, 2022 (7%)

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